Using bioinformatics tools to screen for trypanosomal cathepsin B cysteine protease inhibitors from the SANCDB as a novel therapeutic modality against Human African Trypanosomiasis (HAT)
- Authors: Mokhawa, Gaone
- Date: 2016
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: http://hdl.handle.net/10962/3304 , vital:20470
- Description: Human African Trypanosomiasis (HAT), also known as sleeping sickness, is a fatal chronic disease that is caused by flagellated protozoans, Trypanosoma brucei gambiense and Trypanosoma brucei rhodesiense. HAT is spread by a bite from an infected tsetse fly of the Glosina genus. Up to 60 million people in 36 countries in sub-Saharan Africa are at a risk of infection from HAT with up to 30 000 deaths reported every year. Current chemotherapy for HAT is insufficient since the available drugs exhibit unacceptable side effects (toxicity) and parasite resistance. Novel treatments and approaches for development of specific and more potent drugs for HAT are therefore required. One approach is to target vital proteins that are essential to the life cycle of the parasite. The main interest of this study is to explore Trypanosoma brucei cathepsin B-like protease (TbCatB) structural and functional properties with the primary goal of discovering non peptide small molecule inhibitors of TbCatB using bioinformatics approaches. TbCatB is a papain family C1 cysteine protease which belongs to clan CA group and it has emerged as a potential HAT drug target. Papain family cysteine proteases of Clan CA group of Trypanosoma brucei (rhodesain and TbCatB) have demonstrated potential as chemotherapeutic targets using synthetic protease inhibitors like Z-Phe-Ala-CHN2 to kill the parasite in vitro and in vivo. TbCatB has been identified as the essential cysteine protease of T. brucei since mRNA silencing of TbCatB killed the parasite and resulted in a cure in mice infected with T. brucei while mRNA silencing of rhodesain only extended mice life. TbCatB is therefore a promising drug target against HAT and the discovery and development of compounds that can selectively inhibit TbCatB without posing any danger to the human host represent a great therapeutic solution for treatment of HAT. To understand protein-inhibitor interactions, useful information can be obtained from high resolution protease-inhibitor crystal structure complexes. This study aims to use bioinformatics approaches to carry out comparative sequence, structural and functional analysis of TbCatB protease and its homologs from T. congolense, T, cruzi, T. vivax and H. sapien as well as to identify non-peptide small molecule inhibitors of TbCatB cysteine proteases from natural compounds of South African origin. Sequences of TbCatB (PDB ID: 3HHI) homologs were retrieved by a BLAST search. Human cathepsin B (PDB ID: 3CBJ) was selected from a list of templates for homology modelling found by HHpred. MODELLER version 9.10 program was used to generate a hundred models for T. congolense, T, cruzi and T. vivax cathepsin B like proteases using 3HHI and 3CBJ as templates. The best models were chosen based on their low DOPE Z scores before validation using MetaMQAPII, ANOLEA, PROCHECK and QMEAN6. The DOPE Z scores and the RMSD (RMS) values of the calculated models indicate that the models are of acceptable energy (stability) and fold (conformation). Results from the different MQAPs indicate the models are of acceptable quality and they can be used for docking studies. High throughput screening of SANCDB using AutoDock Vina revealed nine compounds, SANC00 478, 479, 480, 481, 482, 488, 489, 490 and 491, having a strong affinity for Trypanosoma spp. cathepsin B proteases than HsCatB. SANC00488 has the strongest binding to Trypanosoma spp. cathepsin B proteases and the weakest binding to HsCatB protease. Molecular dynamics (MD) simulations show that the complexes between SANC00488 and TbCatB, TcCatB, TcrCatB and TvCatB are stable and do not come apart during simulation. The complex between this compound and HsCatB however is unstable and comes apart during simulation. Residues that are important for the stability of SANC00488-TbCatB complex are Gly328 of the S2 subsite, Phe208, and Ala256. In conclusion SANC00488 is a good candidate for development of a drug against HAT.
- Full Text:
- Date Issued: 2016
- Authors: Mokhawa, Gaone
- Date: 2016
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: http://hdl.handle.net/10962/3304 , vital:20470
- Description: Human African Trypanosomiasis (HAT), also known as sleeping sickness, is a fatal chronic disease that is caused by flagellated protozoans, Trypanosoma brucei gambiense and Trypanosoma brucei rhodesiense. HAT is spread by a bite from an infected tsetse fly of the Glosina genus. Up to 60 million people in 36 countries in sub-Saharan Africa are at a risk of infection from HAT with up to 30 000 deaths reported every year. Current chemotherapy for HAT is insufficient since the available drugs exhibit unacceptable side effects (toxicity) and parasite resistance. Novel treatments and approaches for development of specific and more potent drugs for HAT are therefore required. One approach is to target vital proteins that are essential to the life cycle of the parasite. The main interest of this study is to explore Trypanosoma brucei cathepsin B-like protease (TbCatB) structural and functional properties with the primary goal of discovering non peptide small molecule inhibitors of TbCatB using bioinformatics approaches. TbCatB is a papain family C1 cysteine protease which belongs to clan CA group and it has emerged as a potential HAT drug target. Papain family cysteine proteases of Clan CA group of Trypanosoma brucei (rhodesain and TbCatB) have demonstrated potential as chemotherapeutic targets using synthetic protease inhibitors like Z-Phe-Ala-CHN2 to kill the parasite in vitro and in vivo. TbCatB has been identified as the essential cysteine protease of T. brucei since mRNA silencing of TbCatB killed the parasite and resulted in a cure in mice infected with T. brucei while mRNA silencing of rhodesain only extended mice life. TbCatB is therefore a promising drug target against HAT and the discovery and development of compounds that can selectively inhibit TbCatB without posing any danger to the human host represent a great therapeutic solution for treatment of HAT. To understand protein-inhibitor interactions, useful information can be obtained from high resolution protease-inhibitor crystal structure complexes. This study aims to use bioinformatics approaches to carry out comparative sequence, structural and functional analysis of TbCatB protease and its homologs from T. congolense, T, cruzi, T. vivax and H. sapien as well as to identify non-peptide small molecule inhibitors of TbCatB cysteine proteases from natural compounds of South African origin. Sequences of TbCatB (PDB ID: 3HHI) homologs were retrieved by a BLAST search. Human cathepsin B (PDB ID: 3CBJ) was selected from a list of templates for homology modelling found by HHpred. MODELLER version 9.10 program was used to generate a hundred models for T. congolense, T, cruzi and T. vivax cathepsin B like proteases using 3HHI and 3CBJ as templates. The best models were chosen based on their low DOPE Z scores before validation using MetaMQAPII, ANOLEA, PROCHECK and QMEAN6. The DOPE Z scores and the RMSD (RMS) values of the calculated models indicate that the models are of acceptable energy (stability) and fold (conformation). Results from the different MQAPs indicate the models are of acceptable quality and they can be used for docking studies. High throughput screening of SANCDB using AutoDock Vina revealed nine compounds, SANC00 478, 479, 480, 481, 482, 488, 489, 490 and 491, having a strong affinity for Trypanosoma spp. cathepsin B proteases than HsCatB. SANC00488 has the strongest binding to Trypanosoma spp. cathepsin B proteases and the weakest binding to HsCatB protease. Molecular dynamics (MD) simulations show that the complexes between SANC00488 and TbCatB, TcCatB, TcrCatB and TvCatB are stable and do not come apart during simulation. The complex between this compound and HsCatB however is unstable and comes apart during simulation. Residues that are important for the stability of SANC00488-TbCatB complex are Gly328 of the S2 subsite, Phe208, and Ala256. In conclusion SANC00488 is a good candidate for development of a drug against HAT.
- Full Text:
- Date Issued: 2016
Application of computational methods in elucidating the isomerization step in the biosynthesis of coumarins
- Authors: Tshiwawa, Tendamudzimu
- Date: 2019
- Subjects: Coumarins , Isomerization , Biosynthesis , Organic compounds -- Synthesis , Cinnamic acid
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/67646 , vital:29124
- Description: The identity of the enzyme(s) responsible for the biosynthetic transformation of cinnamic acid derivatives to important, naturally occurring coumarins has yet to be established. This study constitutes a high-level theoretical analysis of the possibility that a recently reported molecular mechanism of the synthesis of coumarins from Baylis-Hillman adducts, may provide a viable model for three critical phases in the biosynthetic pathway Particular attention has been given to the first of these phases: i) E→Z isomerisation of the cinnamic acid precursor; ii) Cyclisation (lactonisation) to the hemi-acetal intermediate; and ii) Dehydration to afford the coumarin derivative. In order to accomplish this analysis, an enzyme capable, theoretically, of effecting this E→Z isomerisation required identification, and its potential involvement in the transformation mechanism explored. Combined Molecular Mechanics and high-level Quantum Mechanical/DFT calculations were used to access complementary models of appropriate complexes and relevant processes within the enzyme active sites of a range of eleven Chalcone Isomerase (CHI) enzyme candidates, the structures of which were downloaded from the Protein Data Bank. Detailed B3LYP/6-31+G(d,p) calculations have provided pictures of the relative populations of conformations within the ensemble of conformations available at normal temperatures. Conformations of several protonation states of cinnamic acid derivatives have been studied in this way, and the results obtained showed that coupled protonation and deprotonation of (E)-o-coumaric acid provides a viable approach to achieve the E→Z isomerization. In silico docking of the B3LYP/6-31+G(d,p) optimized (E)-o-coumaric acid derivatives in the active sites of each of the candidate CHI enzymes (CHI) revealed that (E)-o-coumaric acid fits well within the active sites of Medicago Sativa CHI crystallographic structures with 1FM8 showing best potential for not only accommodating (E)-o-coumaric acid , but also providing appropriate protein active site residues to effect the simultaneous protonation and deprotonation of the substrate , two residues being optimally placed to facilitate these critical processes. Further exploration of the chemical properties and qualities of selected CHI enzymes, undertaken using High Throughput Virtual Screening (HTVS), confirmed 1FM8 as a viable choice for further studies of the enzyme-catalysed E→Z isomerization of (E)-o-coumaric acid. A molecular dynamics study, performed to further evaluate the evolution of (E)-o-coumaric acid in the CHI active site over time, showed that the ligand in the 1FM8 active site is not only stable, but also that the desired protein-ligand interactions persist throughout the simulation period to facilitate the E→Z isomerization. An integrated molecular orbital and molecular mechanics (ONIOM) study of the 1FM8-(E)-o-coumaric acid complex, involving the direct protonation and deprotonation of the ligand by protein residues; has provided a plausible mechanism for the E → Z isomerization of (E)-o-coumaric acid within the 1FM8 active site; a transition state complex (with an activation energy of ca. 50 kCal.mol-1) has been located and its connection with both the (E)- and (Z)-o-coumaric acid isomer has been confirmed by Intrinsic Reaction Coordinate (IRC) calculations. More realistic models of the 1FM8-(E)-o-coumaric acid complex, with the inclusion of water solvent molecules, have been obtained at both the QM/MM and adaptive QM/MM levels which simulate the dynamic active site at the QM level. The results indicate that the simultaneous protonation and deprotonation of (E)-o-coumaric acid within the CHI enzyme is a water-mediated process – a conclusion consistent with similar reported processes. Visual inspection of the 1FM8-(Z)-o-coumaric acid complex reveals both the necessary orientation of the phenolic and carboxylic acid moieties of the (Z)-o-coumaric acid and the presence of appropriate, proximal active site residues with the potential to permit catalysis of the subsequent lactonisation and dehydration steps required to generate coumarin.
- Full Text:
- Date Issued: 2019
- Authors: Tshiwawa, Tendamudzimu
- Date: 2019
- Subjects: Coumarins , Isomerization , Biosynthesis , Organic compounds -- Synthesis , Cinnamic acid
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/67646 , vital:29124
- Description: The identity of the enzyme(s) responsible for the biosynthetic transformation of cinnamic acid derivatives to important, naturally occurring coumarins has yet to be established. This study constitutes a high-level theoretical analysis of the possibility that a recently reported molecular mechanism of the synthesis of coumarins from Baylis-Hillman adducts, may provide a viable model for three critical phases in the biosynthetic pathway Particular attention has been given to the first of these phases: i) E→Z isomerisation of the cinnamic acid precursor; ii) Cyclisation (lactonisation) to the hemi-acetal intermediate; and ii) Dehydration to afford the coumarin derivative. In order to accomplish this analysis, an enzyme capable, theoretically, of effecting this E→Z isomerisation required identification, and its potential involvement in the transformation mechanism explored. Combined Molecular Mechanics and high-level Quantum Mechanical/DFT calculations were used to access complementary models of appropriate complexes and relevant processes within the enzyme active sites of a range of eleven Chalcone Isomerase (CHI) enzyme candidates, the structures of which were downloaded from the Protein Data Bank. Detailed B3LYP/6-31+G(d,p) calculations have provided pictures of the relative populations of conformations within the ensemble of conformations available at normal temperatures. Conformations of several protonation states of cinnamic acid derivatives have been studied in this way, and the results obtained showed that coupled protonation and deprotonation of (E)-o-coumaric acid provides a viable approach to achieve the E→Z isomerization. In silico docking of the B3LYP/6-31+G(d,p) optimized (E)-o-coumaric acid derivatives in the active sites of each of the candidate CHI enzymes (CHI) revealed that (E)-o-coumaric acid fits well within the active sites of Medicago Sativa CHI crystallographic structures with 1FM8 showing best potential for not only accommodating (E)-o-coumaric acid , but also providing appropriate protein active site residues to effect the simultaneous protonation and deprotonation of the substrate , two residues being optimally placed to facilitate these critical processes. Further exploration of the chemical properties and qualities of selected CHI enzymes, undertaken using High Throughput Virtual Screening (HTVS), confirmed 1FM8 as a viable choice for further studies of the enzyme-catalysed E→Z isomerization of (E)-o-coumaric acid. A molecular dynamics study, performed to further evaluate the evolution of (E)-o-coumaric acid in the CHI active site over time, showed that the ligand in the 1FM8 active site is not only stable, but also that the desired protein-ligand interactions persist throughout the simulation period to facilitate the E→Z isomerization. An integrated molecular orbital and molecular mechanics (ONIOM) study of the 1FM8-(E)-o-coumaric acid complex, involving the direct protonation and deprotonation of the ligand by protein residues; has provided a plausible mechanism for the E → Z isomerization of (E)-o-coumaric acid within the 1FM8 active site; a transition state complex (with an activation energy of ca. 50 kCal.mol-1) has been located and its connection with both the (E)- and (Z)-o-coumaric acid isomer has been confirmed by Intrinsic Reaction Coordinate (IRC) calculations. More realistic models of the 1FM8-(E)-o-coumaric acid complex, with the inclusion of water solvent molecules, have been obtained at both the QM/MM and adaptive QM/MM levels which simulate the dynamic active site at the QM level. The results indicate that the simultaneous protonation and deprotonation of (E)-o-coumaric acid within the CHI enzyme is a water-mediated process – a conclusion consistent with similar reported processes. Visual inspection of the 1FM8-(Z)-o-coumaric acid complex reveals both the necessary orientation of the phenolic and carboxylic acid moieties of the (Z)-o-coumaric acid and the presence of appropriate, proximal active site residues with the potential to permit catalysis of the subsequent lactonisation and dehydration steps required to generate coumarin.
- Full Text:
- Date Issued: 2019
Falcipains as malarial drug targets
- Authors: Kanzi, Aquillah Mumo
- Date: 2013
- Subjects: Malaria Malaria -- Chemotherapy Plasmodium falciparum Antimalarials -- Development Cysteine proteinases Cysteine proteinases -- Inhibitors Papain Drug development Bioinformatics
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3897 , http://hdl.handle.net/10962/d1003842
- Description: Malaria is an infectious disease caused by parasites of the Plasmodium genus with mortality rates of more than a million annually, hence a major global public health concern. Plasmodium falciparum (P. falciparum) accounts for over 90% of malaria incidence. Increased resistance to antimalarial drugs by the Plasmodium parasite, coupled with the lack of an effective malaria vaccine necessitates the urgent need for new research avenues to develop novel and more potent antimalarial drugs. This study focused on falcipains, a group of P. falciparum cysteine proteases that belong to the clan CA and papain family C1, that have emerged as potential drug targets due to their involvement in a range of crucial functions in the P. falciparum life cycle. Recently, falcipain-2 has been validated as a drug target but little is known of its Plasmodium orthologs. Currently, there are several falcipain inhibitors that have been identified, most of which are peptide based but none has proceeded to drug development due to associated poor pharmacological profiles and susceptibility to degradation by host cysteine proteases. Non-peptides inhibitors have been shown to be more stable in vivo but limited information exists. In vivo studies on falcipain-2 and falcipain-3 inhibitors have also been complicated by varying outcomes, thus a good understanding of the structural variations of falcipain Plasmodium orthologs at the active site could go a long way to ease in vivo results interpretation and effective inhibitor design. In this study, we use bioinformatics approaches to perform comparative sequence and structural analysis and molecular docking to characterize protein-inhibitor interactions of falcipain homologs at the active site. Known FP-2 and FP-3 small molecule nonpeptide inhibitors were used to identify residue variations and their effect on inhibitor binding. This was done with the aim of screening a collection of selected non-peptide compounds of South African natural origin to identify possible new inhibitor leads. Natural compounds with high binding affinities across all Plasmodium orthologs were identified. These compounds were then used to search the ZINC database for similar compounds which could have better binding affinities across all selected falcipain homologs. Compounds with high binding affinities across all Plasmodium orthologs were found.
- Full Text:
- Date Issued: 2013
- Authors: Kanzi, Aquillah Mumo
- Date: 2013
- Subjects: Malaria Malaria -- Chemotherapy Plasmodium falciparum Antimalarials -- Development Cysteine proteinases Cysteine proteinases -- Inhibitors Papain Drug development Bioinformatics
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:3897 , http://hdl.handle.net/10962/d1003842
- Description: Malaria is an infectious disease caused by parasites of the Plasmodium genus with mortality rates of more than a million annually, hence a major global public health concern. Plasmodium falciparum (P. falciparum) accounts for over 90% of malaria incidence. Increased resistance to antimalarial drugs by the Plasmodium parasite, coupled with the lack of an effective malaria vaccine necessitates the urgent need for new research avenues to develop novel and more potent antimalarial drugs. This study focused on falcipains, a group of P. falciparum cysteine proteases that belong to the clan CA and papain family C1, that have emerged as potential drug targets due to their involvement in a range of crucial functions in the P. falciparum life cycle. Recently, falcipain-2 has been validated as a drug target but little is known of its Plasmodium orthologs. Currently, there are several falcipain inhibitors that have been identified, most of which are peptide based but none has proceeded to drug development due to associated poor pharmacological profiles and susceptibility to degradation by host cysteine proteases. Non-peptides inhibitors have been shown to be more stable in vivo but limited information exists. In vivo studies on falcipain-2 and falcipain-3 inhibitors have also been complicated by varying outcomes, thus a good understanding of the structural variations of falcipain Plasmodium orthologs at the active site could go a long way to ease in vivo results interpretation and effective inhibitor design. In this study, we use bioinformatics approaches to perform comparative sequence and structural analysis and molecular docking to characterize protein-inhibitor interactions of falcipain homologs at the active site. Known FP-2 and FP-3 small molecule nonpeptide inhibitors were used to identify residue variations and their effect on inhibitor binding. This was done with the aim of screening a collection of selected non-peptide compounds of South African natural origin to identify possible new inhibitor leads. Natural compounds with high binding affinities across all Plasmodium orthologs were identified. These compounds were then used to search the ZINC database for similar compounds which could have better binding affinities across all selected falcipain homologs. Compounds with high binding affinities across all Plasmodium orthologs were found.
- Full Text:
- Date Issued: 2013
Application of computer-aided drug design for identification of P. falciparum inhibitors
- Authors: Diallo, Bakary N’tji
- Date: 2021-10-29
- Subjects: Plasmodium falciparum , Malaria -- Chemotherapy , Molecular dynamics , Antimalarials , Cheminformatics , Drug development , Ligand binding (Biochemistry) , Plasmodium falciparum1-deoxy-D-xylulose-5-phosphate reductoisomerase (PfDXR) , South African Natural Compounds Database
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/192798 , vital:45265 , 10.21504/10962/192798
- Description: Malaria is a millennia-old disease with the first recorded cases dating back to 2700 BC found in Chinese medical records, and later in other civilizations. It has claimed human lives to such an extent that there are a notable associated socio-economic consequences. Currently, according to the World Health Organization (WHO), Africa holds the highest disease burden with 94% of deaths and 82% of cases with P. falciparum having ~100% prevalence. Chemotherapy, such as artemisinin combination therapy, has been and continues to be the work horse in the fight against the disease, together with seasonal malaria chemoprevention and the use of insecticides. Natural products such as quinine and artemisinin are particularly important in terms of their antimalarial activity. The emphasis in current chemotherapy research is the need for time and cost-effective workflows focussed on new mechanisms of action (MoAs) covering the target candidate profiles (TCPs). Despite a decline in cases over the past decades with, countries increasingly becoming certified malaria free, a stalling trend has been observed in the past five years resulting in missing the 2020 Global Technical Strategy (GTS) milestones. With no effective vaccine, a reduction in funding, slower drug approval than resistance emergence from resistant and invasive vectors, and threats in diagnosis with the pfhrp2/3 gene deletion, malaria remains a major health concern. Motivated by these reasons, the primary aim of this work was a contribution to the antimalarial pipeline through in silico approaches focusing on P. falciparum. We first intended an exploration of malarial targets through a proteome scale screening on 36 targets using multiple metrics to account for the multi-objective nature of drug discovery. The continuous growth of structural data offers the ideal scenario for mining new MoAs covering antimalarials TCPs. This was combined with a repurposing strategy using a set of orally available FDA approved drugs. Further, use was made of time- and cost-effective strategies combining QVina-W efficiency metrics that integrate molecular properties, GRIM rescoring for molecular interactions and a hydrogen mass repartitioning (HMR) molecular dynamics (MD) scheme for accelerated development of antimalarials in the context of resistance. This pipeline further integrates a complex ranking for better drug-target selectivity, and normalization strategies to overcome docking scoring function bias. The different metrics, ranking, normalization strategies and their combinations were first assessed using their mean ranking error (MRE). A version combining all metrics was used to select 36 unique protein-ligand complexes, assessed in MD, with the final retention of 25. From the 16 in vitro tested hits of the 25, fingolimod, abiraterone, prazosin, and terazosin showed antiplasmodial activity with IC50 2.21, 3.37, 16.67 and 34.72 μM respectively and of these, only fingolimod was found to be not safe with respect to human cell viability. These compounds were predicted active on different molecular targets, abiraterone was predicted to interact with a putative liver-stage essential target, hence promising as a transmission-blocking agent. The pipeline had a promising 25% hit rate considering the proteome-scale and use of cost-effective approaches. Secondly, we focused on Plasmodium falciparum 1-deoxy-D-xylulose-5-phosphate reductoisomerase (PfDXR) using a more extensive screening pipeline to overcome some of the current in silico screening limitations. Starting from the ZINC lead-like library of ~3M, hierarchical ligand-based virtual screening (LBVS) and structure-based virtual screening (SBVS) approaches with molecular docking and re-scoring using eleven scoring functions (SFs) were used. Later ranking with an exponential consensus strategy was included. Selected hits were further assessed through Molecular Mechanics Poisson-Boltzmann Surface Area (MM-PBSA), advanced MD sampling in a ligand pulling simulations and (Weighted Histogram Analysis Method) WHAM analysis for umbrella sampling (US) to derive binding free energies. Four leads had better predicted affinities in US than LC5, a 280 nM potent PfDXR inhibitor with ZINC000050633276 showing a promising binding of -20.43 kcal/mol. As shown with fosmidomycin, DXR inhibition offers fast acting compounds fulfilling antimalarials TCP1. Yet, fosmidomycin has a high polarity causing its short half-life and hampering its clinical use. These leads scaffolds are different from fosmidomycin and hence may offer better pharmacokinetic and pharmacodynamic properties and may also be promising for lead optimization. A combined analysis of residues’ contributions to the free energy of binding in MM-PBSA and to steered molecular dynamics (SMD) Fmax indicated GLU233, CYS268, SER270, TRP296, and HIS341 as exploitable for compound optimization. Finally, we updated the SANCDB library with new NPs and their commercially available analogs as a solution to NP availability. The library is extended to 1005 compounds from its initial 600 compounds and the database is integrated to Mcule and Molport APIs for analogs automatic update. The new set may contribute to virtual screening and to antimalarials as the most effective ones have NP origin. , Thesis (PhD) -- Faculty of Science, Biochemistry and Microbiology, 2021
- Full Text:
- Date Issued: 2021-10-29
- Authors: Diallo, Bakary N’tji
- Date: 2021-10-29
- Subjects: Plasmodium falciparum , Malaria -- Chemotherapy , Molecular dynamics , Antimalarials , Cheminformatics , Drug development , Ligand binding (Biochemistry) , Plasmodium falciparum1-deoxy-D-xylulose-5-phosphate reductoisomerase (PfDXR) , South African Natural Compounds Database
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/192798 , vital:45265 , 10.21504/10962/192798
- Description: Malaria is a millennia-old disease with the first recorded cases dating back to 2700 BC found in Chinese medical records, and later in other civilizations. It has claimed human lives to such an extent that there are a notable associated socio-economic consequences. Currently, according to the World Health Organization (WHO), Africa holds the highest disease burden with 94% of deaths and 82% of cases with P. falciparum having ~100% prevalence. Chemotherapy, such as artemisinin combination therapy, has been and continues to be the work horse in the fight against the disease, together with seasonal malaria chemoprevention and the use of insecticides. Natural products such as quinine and artemisinin are particularly important in terms of their antimalarial activity. The emphasis in current chemotherapy research is the need for time and cost-effective workflows focussed on new mechanisms of action (MoAs) covering the target candidate profiles (TCPs). Despite a decline in cases over the past decades with, countries increasingly becoming certified malaria free, a stalling trend has been observed in the past five years resulting in missing the 2020 Global Technical Strategy (GTS) milestones. With no effective vaccine, a reduction in funding, slower drug approval than resistance emergence from resistant and invasive vectors, and threats in diagnosis with the pfhrp2/3 gene deletion, malaria remains a major health concern. Motivated by these reasons, the primary aim of this work was a contribution to the antimalarial pipeline through in silico approaches focusing on P. falciparum. We first intended an exploration of malarial targets through a proteome scale screening on 36 targets using multiple metrics to account for the multi-objective nature of drug discovery. The continuous growth of structural data offers the ideal scenario for mining new MoAs covering antimalarials TCPs. This was combined with a repurposing strategy using a set of orally available FDA approved drugs. Further, use was made of time- and cost-effective strategies combining QVina-W efficiency metrics that integrate molecular properties, GRIM rescoring for molecular interactions and a hydrogen mass repartitioning (HMR) molecular dynamics (MD) scheme for accelerated development of antimalarials in the context of resistance. This pipeline further integrates a complex ranking for better drug-target selectivity, and normalization strategies to overcome docking scoring function bias. The different metrics, ranking, normalization strategies and their combinations were first assessed using their mean ranking error (MRE). A version combining all metrics was used to select 36 unique protein-ligand complexes, assessed in MD, with the final retention of 25. From the 16 in vitro tested hits of the 25, fingolimod, abiraterone, prazosin, and terazosin showed antiplasmodial activity with IC50 2.21, 3.37, 16.67 and 34.72 μM respectively and of these, only fingolimod was found to be not safe with respect to human cell viability. These compounds were predicted active on different molecular targets, abiraterone was predicted to interact with a putative liver-stage essential target, hence promising as a transmission-blocking agent. The pipeline had a promising 25% hit rate considering the proteome-scale and use of cost-effective approaches. Secondly, we focused on Plasmodium falciparum 1-deoxy-D-xylulose-5-phosphate reductoisomerase (PfDXR) using a more extensive screening pipeline to overcome some of the current in silico screening limitations. Starting from the ZINC lead-like library of ~3M, hierarchical ligand-based virtual screening (LBVS) and structure-based virtual screening (SBVS) approaches with molecular docking and re-scoring using eleven scoring functions (SFs) were used. Later ranking with an exponential consensus strategy was included. Selected hits were further assessed through Molecular Mechanics Poisson-Boltzmann Surface Area (MM-PBSA), advanced MD sampling in a ligand pulling simulations and (Weighted Histogram Analysis Method) WHAM analysis for umbrella sampling (US) to derive binding free energies. Four leads had better predicted affinities in US than LC5, a 280 nM potent PfDXR inhibitor with ZINC000050633276 showing a promising binding of -20.43 kcal/mol. As shown with fosmidomycin, DXR inhibition offers fast acting compounds fulfilling antimalarials TCP1. Yet, fosmidomycin has a high polarity causing its short half-life and hampering its clinical use. These leads scaffolds are different from fosmidomycin and hence may offer better pharmacokinetic and pharmacodynamic properties and may also be promising for lead optimization. A combined analysis of residues’ contributions to the free energy of binding in MM-PBSA and to steered molecular dynamics (SMD) Fmax indicated GLU233, CYS268, SER270, TRP296, and HIS341 as exploitable for compound optimization. Finally, we updated the SANCDB library with new NPs and their commercially available analogs as a solution to NP availability. The library is extended to 1005 compounds from its initial 600 compounds and the database is integrated to Mcule and Molport APIs for analogs automatic update. The new set may contribute to virtual screening and to antimalarials as the most effective ones have NP origin. , Thesis (PhD) -- Faculty of Science, Biochemistry and Microbiology, 2021
- Full Text:
- Date Issued: 2021-10-29
In silico study of Plasmodium 1-deoxy-dxylulose 5-phosphate reductoisomerase (DXR) for identification of novel inhibitors from SANCDB
- Authors: Diallo, Bakary N'tji
- Date: 2018
- Subjects: Plasmodium 1-deoxy-dxylulose 5-phosphate reductoisomerase , Isoprenoids , Plasmodium , Antimalarials , Malaria -- Chemotherapy , Molecules -- Models , Molecular dynamics , South African Natural Compounds Database
- Language: English
- Type: text , Thesis , Masters , MSc
- Identifier: http://hdl.handle.net/10962/64012 , vital:28523
- Description: Malaria remains a major health concern with a complex parasite constantly developing resistance to the different drugs introduced to treat it, threatening the efficacy of the current ACT treatment recommended by WHO (World Health Organization). Different antimalarial compounds with different mechanisms of action are ideal as this decreases chances of resistance occurring. Inhibiting DXR and consequently the MEP pathway is a good strategy to find a new antimalarial with a novel mode of action. From literature, all the enzymes of the MEP pathway have also been shown to be indispensable for the synthesis of isoprenoids. They have been validated as drug targets and the X-ray structure of each of the enzymes has been solved. DXR is a protein which catalyses the second step of the MEP pathway. There are currently 255 DXR inhibitors in the Binding Database (accessed November 2017) generally based on the fosmidomycin structural scaffold and thus often showing poor drug likeness properties. This study aims to research new DXR inhibitors using in silico techniques. We analysed the protein sequence and built 3D models in close and open conformations for the different Plasmodium sequences. Then SANCDB compounds were screened to identify new potential DXR inhibitors with new chemical scaffolds. Finally, the identified hits were submitted to molecular dynamics studies, preceded by a parameterization of the manganese atom in the protein active site.
- Full Text:
- Date Issued: 2018
- Authors: Diallo, Bakary N'tji
- Date: 2018
- Subjects: Plasmodium 1-deoxy-dxylulose 5-phosphate reductoisomerase , Isoprenoids , Plasmodium , Antimalarials , Malaria -- Chemotherapy , Molecules -- Models , Molecular dynamics , South African Natural Compounds Database
- Language: English
- Type: text , Thesis , Masters , MSc
- Identifier: http://hdl.handle.net/10962/64012 , vital:28523
- Description: Malaria remains a major health concern with a complex parasite constantly developing resistance to the different drugs introduced to treat it, threatening the efficacy of the current ACT treatment recommended by WHO (World Health Organization). Different antimalarial compounds with different mechanisms of action are ideal as this decreases chances of resistance occurring. Inhibiting DXR and consequently the MEP pathway is a good strategy to find a new antimalarial with a novel mode of action. From literature, all the enzymes of the MEP pathway have also been shown to be indispensable for the synthesis of isoprenoids. They have been validated as drug targets and the X-ray structure of each of the enzymes has been solved. DXR is a protein which catalyses the second step of the MEP pathway. There are currently 255 DXR inhibitors in the Binding Database (accessed November 2017) generally based on the fosmidomycin structural scaffold and thus often showing poor drug likeness properties. This study aims to research new DXR inhibitors using in silico techniques. We analysed the protein sequence and built 3D models in close and open conformations for the different Plasmodium sequences. Then SANCDB compounds were screened to identify new potential DXR inhibitors with new chemical scaffolds. Finally, the identified hits were submitted to molecular dynamics studies, preceded by a parameterization of the manganese atom in the protein active site.
- Full Text:
- Date Issued: 2018
An in-silico study of the type II NADH: Quinone Oxidoreductase (ndh2). A new anti-malaria drug target
- Authors: Baye, Bertha Cinthia
- Date: 2022-10-14
- Subjects: Malaria , Plasmodium , Molecular dynamics , Computer simulation , Quinone , Antimalarials , Molecules Models , Docking , Drugs Computer-aided design
- Language: English
- Type: Academic theses , Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/365633 , vital:65767 , DOI https://doi.org/10.21504/10962/365633
- Description: Malaria is caused by Plasmodium parasites, spread to people through the bites of infected female Anopheles mosquitoes. This study focuses on all 5 (Plasmodium falciparum, Plasmodium knowlesi, Plasmodium malariae, Plasmodium ovale and Plasmodium vivax) parasites that cause malaria in humans. Africa is a developing continent, and it is the most affected with an estimation of 90% of more than 400 000 malaria-related deaths reported by the World Health Organization (WHO) report in 2020, in which 61% of that number are children under the ages of five. Malaria resistance was initially observed in early 1986 and with the progression of time anti-malarial drug resistance has only increased. As a result, there is a need to study the malarial proteins mechanism of action and identify alternative treatment strategies for this disease. Type II NADH: quinone oxidoreductase (NDH2) is a monotopic protein that catalyses the electron transfer from NADH to quinone via FAD without a proton-pumping activity, and functions as an initial enzyme, either in addition to or as an alternative to proton-pumping NADH dehydrogenase (complex I) in the respiratory chain of bacteria, archaea, and fungal and plant mitochondrial. The structures for the Plasmodium knowlesi, Plasmodium malariae, Plasmodium ovale and Plasmodium vivax were modelled from the crystal structure of Plasmodium falciparum (5JWA). Compounds from the South African natural compounds database (SANCDB) were docked against both the NDH2 crystal structure and modelled structures. By performing in silico screening the study aimed to find potential compounds that might interrupt the electron transfer to quinone therefore disturbing the enzyme‟s function and thereby possibly eliminating the plasmodium parasite. CHARMM-GUI was used to create the membrane (since this work is with membrane-bound proteins) and to orient the protein on the membrane using OPM server guidelines, the interface produced GROMACS topology files that were used in molecular dynamics simulations. Molecular dynamics simulations were performed in the Centre for high performance computing (CHPC) cluster under the CHEM0802 project and the trajectories produced were further analysed. In this work not only were hit compounds from SANCDB identified, but also differences in behaviour across species and in the presence or absence of the membrane were described. This highlights the need to include the correct protein environment when studying these systems. , Thesis (PhD) -- Faculty of Science, Biochemistry and Microbiology, 2022
- Full Text:
- Date Issued: 2022-10-14
- Authors: Baye, Bertha Cinthia
- Date: 2022-10-14
- Subjects: Malaria , Plasmodium , Molecular dynamics , Computer simulation , Quinone , Antimalarials , Molecules Models , Docking , Drugs Computer-aided design
- Language: English
- Type: Academic theses , Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/365633 , vital:65767 , DOI https://doi.org/10.21504/10962/365633
- Description: Malaria is caused by Plasmodium parasites, spread to people through the bites of infected female Anopheles mosquitoes. This study focuses on all 5 (Plasmodium falciparum, Plasmodium knowlesi, Plasmodium malariae, Plasmodium ovale and Plasmodium vivax) parasites that cause malaria in humans. Africa is a developing continent, and it is the most affected with an estimation of 90% of more than 400 000 malaria-related deaths reported by the World Health Organization (WHO) report in 2020, in which 61% of that number are children under the ages of five. Malaria resistance was initially observed in early 1986 and with the progression of time anti-malarial drug resistance has only increased. As a result, there is a need to study the malarial proteins mechanism of action and identify alternative treatment strategies for this disease. Type II NADH: quinone oxidoreductase (NDH2) is a monotopic protein that catalyses the electron transfer from NADH to quinone via FAD without a proton-pumping activity, and functions as an initial enzyme, either in addition to or as an alternative to proton-pumping NADH dehydrogenase (complex I) in the respiratory chain of bacteria, archaea, and fungal and plant mitochondrial. The structures for the Plasmodium knowlesi, Plasmodium malariae, Plasmodium ovale and Plasmodium vivax were modelled from the crystal structure of Plasmodium falciparum (5JWA). Compounds from the South African natural compounds database (SANCDB) were docked against both the NDH2 crystal structure and modelled structures. By performing in silico screening the study aimed to find potential compounds that might interrupt the electron transfer to quinone therefore disturbing the enzyme‟s function and thereby possibly eliminating the plasmodium parasite. CHARMM-GUI was used to create the membrane (since this work is with membrane-bound proteins) and to orient the protein on the membrane using OPM server guidelines, the interface produced GROMACS topology files that were used in molecular dynamics simulations. Molecular dynamics simulations were performed in the Centre for high performance computing (CHPC) cluster under the CHEM0802 project and the trajectories produced were further analysed. In this work not only were hit compounds from SANCDB identified, but also differences in behaviour across species and in the presence or absence of the membrane were described. This highlights the need to include the correct protein environment when studying these systems. , Thesis (PhD) -- Faculty of Science, Biochemistry and Microbiology, 2022
- Full Text:
- Date Issued: 2022-10-14
Synthetic analogues of marine bisindole alkaloids as potent selective inhibitors of MRSA pyruvate kinase
- Veale, Clinton Gareth Lancaster
- Authors: Veale, Clinton Gareth Lancaster
- Date: 2014 , 2014-04-02
- Subjects: Alkaloids , Pyruvate kinase , Staphylococcus aureus , Antibiotics , Sponges -- South Africa , Imidazoles , Biological assay , Antibacterial agents
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4563 , http://hdl.handle.net/10962/d1020893
- Description: Globally, methicillin resistant Staphylococcus aureus (MRSA) has become increasingly difficult to manage in the clinic and new antibiotics are required. The structure activity relationship (SAR) study presented in this thesis forms part of an international collaborative effort to identify potent and selective inhibitors of an MRSA pyruvate kinase (PK) enzyme target. In earlier work the known marine natural product bromodeoxytopsentin (1.6), isolated from a South African marine sponge Topsentia pachastrelloides, exhibited selective and significant inhibition of MRSA PK (IC₅₀ 60 nM). Accordingly bromodeoxytopsentin provided the initial chemical scaffold around which our SAR study was developed. Following a comprehensive introduction, providing the necessary background to the research described in subsequent Chapters, this thesis has been divided into three major parts. Part one (Chapter 2) documents the synthesis of two natural imidazole containing topsentin analogues 1.40, 1.46, five new synthetic analogues 1.58—1.61, 2.104. In the process we developed a new method for the synthesis of topsentin derivatives via selenium dioxide mediated oxidation of N-Boc protected 3-acetylindoles to yield glyoxal intermediates which were subsequently cyclized and deprotected to yield the desired products. Interestingly we were able to demonstrate a delicate relationship between the relative equivalents of selenium dioxide and water used during the oxidation step, careful manipulation of which was required to prevent the uncontrolled formation of side products. Synthetic compounds 1.40, 1.46, 1.58—1.61 were found to be potent inhibitors of MRSA PK (IC₅₀ 238, 2.1, 23, 1.4, 6.3 and 3.2 nM respectively) with 1000-10000 fold selectivity for MRSA PK over four human orthologs. In the second part of this thesis (Chapter 3) we report the successful synthesis of a cohort of previously unknown thiazole containing bisindole topsentin analogues 1.62—1.68 via a Hantzsch thiazole synthesis. Bioassay results revealed that these compounds were only moderate inhibitors of MRSA PK (IC₅₀ 5.1—20 μM) which suggested that inhibitory activity was significantly reduced upon substitution of the central imidazole ring of topsentin type analogues with a thiazole type ring. In addition in Chapter 3 we describe unsuccessful attempts to regiospecifically synthesize oxazole and imidazole topsentin analogues through a similar Hantzsch method. As a consequence of our efforts in this regard we investigated three key reactions in depth, namely the synthesis of 2.2, 3.38, 3.40, 3.41 via α-bromination of 3-acetylindole and the synthesis of indolyl-3-carbonylnitriles 2.13, 3.45—3.47 and α-oxo-1H-indole-3-thioacetamides 3.48—3.51. The investigation of the latter led to the isolation and elucidation of two anomalous N,N-dimethyl-1H-indole-3-carboxamides 3.52 and 3.53. Finally the third part of this thesis (Chapter 4) deals with in silico assessment of the binding of both the imidazole and thiazole containing bisindole alkaloids to the MRSA PK protein which initially guided our SAR studies. In this chapter we reveal that there appears to be no correlation between in silico binding predictions and in vitro MRSA PK inhibitory bioassay data. Superficially it seems that binding energy as determined by the docking program used for these studies correlated with the size of the indole substituents and did not reflect IC₅₀ MRSA PK inhibitory data. Although this led us to computationally explore possible alternative binding sites no clear alternative has been identified.
- Full Text:
- Date Issued: 2014
- Authors: Veale, Clinton Gareth Lancaster
- Date: 2014 , 2014-04-02
- Subjects: Alkaloids , Pyruvate kinase , Staphylococcus aureus , Antibiotics , Sponges -- South Africa , Imidazoles , Biological assay , Antibacterial agents
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4563 , http://hdl.handle.net/10962/d1020893
- Description: Globally, methicillin resistant Staphylococcus aureus (MRSA) has become increasingly difficult to manage in the clinic and new antibiotics are required. The structure activity relationship (SAR) study presented in this thesis forms part of an international collaborative effort to identify potent and selective inhibitors of an MRSA pyruvate kinase (PK) enzyme target. In earlier work the known marine natural product bromodeoxytopsentin (1.6), isolated from a South African marine sponge Topsentia pachastrelloides, exhibited selective and significant inhibition of MRSA PK (IC₅₀ 60 nM). Accordingly bromodeoxytopsentin provided the initial chemical scaffold around which our SAR study was developed. Following a comprehensive introduction, providing the necessary background to the research described in subsequent Chapters, this thesis has been divided into three major parts. Part one (Chapter 2) documents the synthesis of two natural imidazole containing topsentin analogues 1.40, 1.46, five new synthetic analogues 1.58—1.61, 2.104. In the process we developed a new method for the synthesis of topsentin derivatives via selenium dioxide mediated oxidation of N-Boc protected 3-acetylindoles to yield glyoxal intermediates which were subsequently cyclized and deprotected to yield the desired products. Interestingly we were able to demonstrate a delicate relationship between the relative equivalents of selenium dioxide and water used during the oxidation step, careful manipulation of which was required to prevent the uncontrolled formation of side products. Synthetic compounds 1.40, 1.46, 1.58—1.61 were found to be potent inhibitors of MRSA PK (IC₅₀ 238, 2.1, 23, 1.4, 6.3 and 3.2 nM respectively) with 1000-10000 fold selectivity for MRSA PK over four human orthologs. In the second part of this thesis (Chapter 3) we report the successful synthesis of a cohort of previously unknown thiazole containing bisindole topsentin analogues 1.62—1.68 via a Hantzsch thiazole synthesis. Bioassay results revealed that these compounds were only moderate inhibitors of MRSA PK (IC₅₀ 5.1—20 μM) which suggested that inhibitory activity was significantly reduced upon substitution of the central imidazole ring of topsentin type analogues with a thiazole type ring. In addition in Chapter 3 we describe unsuccessful attempts to regiospecifically synthesize oxazole and imidazole topsentin analogues through a similar Hantzsch method. As a consequence of our efforts in this regard we investigated three key reactions in depth, namely the synthesis of 2.2, 3.38, 3.40, 3.41 via α-bromination of 3-acetylindole and the synthesis of indolyl-3-carbonylnitriles 2.13, 3.45—3.47 and α-oxo-1H-indole-3-thioacetamides 3.48—3.51. The investigation of the latter led to the isolation and elucidation of two anomalous N,N-dimethyl-1H-indole-3-carboxamides 3.52 and 3.53. Finally the third part of this thesis (Chapter 4) deals with in silico assessment of the binding of both the imidazole and thiazole containing bisindole alkaloids to the MRSA PK protein which initially guided our SAR studies. In this chapter we reveal that there appears to be no correlation between in silico binding predictions and in vitro MRSA PK inhibitory bioassay data. Superficially it seems that binding energy as determined by the docking program used for these studies correlated with the size of the indole substituents and did not reflect IC₅₀ MRSA PK inhibitory data. Although this led us to computationally explore possible alternative binding sites no clear alternative has been identified.
- Full Text:
- Date Issued: 2014
A Comparative study of two copper(II) based metal-organic frameworks : Cu2¼(OH)½B4C•8H2O and Cu2Na(OH)B4C•7H2O
- Authors: Coombes, Matthew
- Date: 2013
- Subjects: Copper , Organometallic compounds , Supramolecular organometallic chemistry
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4533 , http://hdl.handle.net/10962/d1016245
- Description: This study focussed on two copper(II)-containing metal-organic frameworks (MOFs): Cu2Na(OH)B4C•7H2O and Cu2¼(OH)½B4C•8H2O (B4C = 1,2,4,5- benzenetetracarboxylate). They are both covalent, three-dimensional metalorganic framework polymers containing voids filled with water molecules. Both were characterised by elemental analysis, infrared spectroscopy, X-ray powder diffractometry (both in situ and regular), thermogravimetric analysis, differential scanning calorimetry and X-ray photoelectron spectroscopy. These two MOFs are essentially identical, with the only difference being the substitution of sodium by copper at every 4th site (disordered throughout the crystal). The guest inclusion properties of both MOFs were studied and compared. Although both structures collapse on dehydration, it was observed that Cu2Na(OH)B4C•7H2O is able to take up signifcant amounts of water, methanol and ethanol. All these processes are fully reversible. Car-Parrinello molecular dynamics studies suggest that it is a strong interaction between the oxygen atoms on these molecules with the sodium cation of the MOF that is responsible for this signifcant uptake. In contrast, Cu2¼ (OH)½ B4C•8H2O, the MOF without a sodium cation, did not demonstrate any methanol or ethanol uptake, but was able to take up some water. The uptake of water, however, is not a fully reversible process. The absence of sodium likely results in insuffcient energy to draw methanol and ethanol into the framework, while a subtle rotation of a carboxylate group on dehydration decreases the ability of the framework to form hydrogen bonds, thus reducing the ability to take up water. A series of hydrothermal syntheses were performed in order to develop a method of synthesis superior to the current gel-based synthesis that requires several months and has poor yields. The hydrothermal products were characterized by elemental analysis, infrared spectroscopy, X-ray powder diffractometry, thermogravimetric analysis and differential scanning calorimetry. It was shown that the MOF Cu2Na(OH)B4C•7H2O may be synthesised in almost 100% yield by using a temperature of 120°C over a period of 72 hours. It was not possible to synthesise Cu2¼ (OH)½ B4C•8H2O in a 100% yield - it was only obtained as a minor product.
- Full Text:
- Date Issued: 2013
- Authors: Coombes, Matthew
- Date: 2013
- Subjects: Copper , Organometallic compounds , Supramolecular organometallic chemistry
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4533 , http://hdl.handle.net/10962/d1016245
- Description: This study focussed on two copper(II)-containing metal-organic frameworks (MOFs): Cu2Na(OH)B4C•7H2O and Cu2¼(OH)½B4C•8H2O (B4C = 1,2,4,5- benzenetetracarboxylate). They are both covalent, three-dimensional metalorganic framework polymers containing voids filled with water molecules. Both were characterised by elemental analysis, infrared spectroscopy, X-ray powder diffractometry (both in situ and regular), thermogravimetric analysis, differential scanning calorimetry and X-ray photoelectron spectroscopy. These two MOFs are essentially identical, with the only difference being the substitution of sodium by copper at every 4th site (disordered throughout the crystal). The guest inclusion properties of both MOFs were studied and compared. Although both structures collapse on dehydration, it was observed that Cu2Na(OH)B4C•7H2O is able to take up signifcant amounts of water, methanol and ethanol. All these processes are fully reversible. Car-Parrinello molecular dynamics studies suggest that it is a strong interaction between the oxygen atoms on these molecules with the sodium cation of the MOF that is responsible for this signifcant uptake. In contrast, Cu2¼ (OH)½ B4C•8H2O, the MOF without a sodium cation, did not demonstrate any methanol or ethanol uptake, but was able to take up some water. The uptake of water, however, is not a fully reversible process. The absence of sodium likely results in insuffcient energy to draw methanol and ethanol into the framework, while a subtle rotation of a carboxylate group on dehydration decreases the ability of the framework to form hydrogen bonds, thus reducing the ability to take up water. A series of hydrothermal syntheses were performed in order to develop a method of synthesis superior to the current gel-based synthesis that requires several months and has poor yields. The hydrothermal products were characterized by elemental analysis, infrared spectroscopy, X-ray powder diffractometry, thermogravimetric analysis and differential scanning calorimetry. It was shown that the MOF Cu2Na(OH)B4C•7H2O may be synthesised in almost 100% yield by using a temperature of 120°C over a period of 72 hours. It was not possible to synthesise Cu2¼ (OH)½ B4C•8H2O in a 100% yield - it was only obtained as a minor product.
- Full Text:
- Date Issued: 2013
Structural analysis of effects of mutations on HIV-1 subtype C protease active site
- Mathu, Alexander Muchugia Nganga
- Authors: Mathu, Alexander Muchugia Nganga
- Date: 2012
- Subjects: HIV (Viruses) -- Research , HIV infections -- Treatment -- Research , Protease inhibitors -- Research , Viruses -- Effect of drugs on -- Research
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4013 , http://hdl.handle.net/10962/d1004073 , HIV (Viruses) -- Research , HIV infections -- Treatment -- Research , Protease inhibitors -- Research , Viruses -- Effect of drugs on -- Research
- Description: HIV/AIDS is a global pandemic that poses a great threat especially in Sub-Saharan Africa where the highest population of those infected with the virus is found. It has far reaching medical, socio-economic and scientific implications. The HIV-1 protease enzyme is a prime therapeutic target that has been exploited in an effort to reduce morbidity and mortality. However problems arise from drug toxicity and drug-resistant mutations of the protease which is a motivation for research for new, safer and effective therapies. Evidence exists to show that there are significant genomic differences in Subtype B and C that have a negative effect on the intrinsic binding of inhibitors. It is imperative to look at all perspectives from epidemiological, molecular to the pharmacological ones so as to achieve rational design of therapeutic agents. This study involved the use of in silico structural analysis of the effects of mutations in the active site. The data was provided by the National Institute of Communicable Diseases consisting of HIV-1 Subtype C protease sequences of 29 infants exhibiting drug-resistance to ritonavir and lopinavir. The major active site mutations causing drug resistance identified in this study were M46I, I54V and V82A using the Stanford HIV database tool. Homology modeling without extra restraints produced models with improved quality in comparison to those with restraints. MetaMQAPII results differed when models were visualized as dimers giving erroneous modeled regions in comparison to monomers. A broader study with a larger dataset of HIV-1 subtype C protease sequences is required to increase statistical confidence and in order to identify the pattern of drug resistant mutations. Homology modeling without extra restraints is preferred for calculating homology models for the HIV-1 subtype C. Further investigations needs to be done to ascertain the accuracy of validation results for dimers from MetaMQAPII as it is designed for evaluation of monomers.
- Full Text:
- Date Issued: 2012
- Authors: Mathu, Alexander Muchugia Nganga
- Date: 2012
- Subjects: HIV (Viruses) -- Research , HIV infections -- Treatment -- Research , Protease inhibitors -- Research , Viruses -- Effect of drugs on -- Research
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4013 , http://hdl.handle.net/10962/d1004073 , HIV (Viruses) -- Research , HIV infections -- Treatment -- Research , Protease inhibitors -- Research , Viruses -- Effect of drugs on -- Research
- Description: HIV/AIDS is a global pandemic that poses a great threat especially in Sub-Saharan Africa where the highest population of those infected with the virus is found. It has far reaching medical, socio-economic and scientific implications. The HIV-1 protease enzyme is a prime therapeutic target that has been exploited in an effort to reduce morbidity and mortality. However problems arise from drug toxicity and drug-resistant mutations of the protease which is a motivation for research for new, safer and effective therapies. Evidence exists to show that there are significant genomic differences in Subtype B and C that have a negative effect on the intrinsic binding of inhibitors. It is imperative to look at all perspectives from epidemiological, molecular to the pharmacological ones so as to achieve rational design of therapeutic agents. This study involved the use of in silico structural analysis of the effects of mutations in the active site. The data was provided by the National Institute of Communicable Diseases consisting of HIV-1 Subtype C protease sequences of 29 infants exhibiting drug-resistance to ritonavir and lopinavir. The major active site mutations causing drug resistance identified in this study were M46I, I54V and V82A using the Stanford HIV database tool. Homology modeling without extra restraints produced models with improved quality in comparison to those with restraints. MetaMQAPII results differed when models were visualized as dimers giving erroneous modeled regions in comparison to monomers. A broader study with a larger dataset of HIV-1 subtype C protease sequences is required to increase statistical confidence and in order to identify the pattern of drug resistant mutations. Homology modeling without extra restraints is preferred for calculating homology models for the HIV-1 subtype C. Further investigations needs to be done to ascertain the accuracy of validation results for dimers from MetaMQAPII as it is designed for evaluation of monomers.
- Full Text:
- Date Issued: 2012
The investigation of type-specific features of the copper coordinating AA9 proteins and their effect on the interaction with crystalline cellulose using molecular dynamics studies
- Authors: Moses, Vuyani
- Date: 2018
- Subjects: Copper proteins , Cellulose , Molecular dynamics , Cellulose -- Biodegradation , Bioinformatics
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/58327 , vital:27230
- Description: AA9 proteins are metallo-enzymes which are crucial for the early stages of cellulose degradation. AA9 proteins have been suggested to cleave glycosidic bonds linking cellulose through the use of their Cu2+ coordinating active site. AA9 proteins possess different regioselectivities depending on the resulting cleavage they form and as result, are grouped accordingly. Type 1 AA9 proteins cleave the C1 carbon of cellulose while Type 2 AA9 proteins cleave the C4 carbon and Type 3 AA9 proteins cleave either C1 or C4 carbons. The steric congestion of the AA9 active site has been proposed to be a contributor to the observed regioselectivity. As such, a bioinformatics characterisation of type-specific sequence and structural features was performed. Initially AA9 protein sequences were obtained from the Pfam database and multiple sequence alignment was performed. The sequences were phylogenetically characterised and sequences were grouped into their respective types and sub-groups were identified. A selection analysis was performed on AA9 LPMO types to determine the selective pressure acting on AA9 protein residues. Motif discovery was then performed to identify conserved sequence motifs in AA9 proteins. Once type-specific sequence features were identified structural mapping was performed to assess possible effects on substrate interaction. Physicochemical property analysis was also performed to assess biochemical differences between AA9 LPMO types. Molecular dynamics (MD) simulations were then employed to dynamically assess the consequences of the discovered type-specific features on AA9-cellulose interaction. Due to the absence of AA9 specific force field parameters MD simulations were not readily applicable. As a result, Potential Energy Surface (PES) scans were performed to evaluate the force field parameters for the AA9 active site using the PM6 semi empirical approach and least squares fitting. A Type 1 AA9 active site was constructed from the crystal structure 4B5Q, encompassing only the Cu2+ coordinating residues, the Cu2+ ion and two water residues. Due to the similarity in AA9 active sites, the Type force field parameters were validated on all three AA9 LPMO types. Two MD simulations for each AA9 LPMO types were conducted using two separate Lennard-Jones parameter sets. Once completed, the MD trajectories were analysed for various features including the RMSD, RMSF, radius of gyration, coordination during simulation, hydrogen bonding, secondary structure conservation and overall protein movement. Force field parameters were successfully evaluated and validated for AA9 proteins. MD simulations of AA9 proteins were able to reveal the presence of unique type-specific binding modes of AA9 active sites to cellulose. These binding modes were characterised by the presence of unique type-specific loops which were present in Type 2 and 3 AA9 proteins but not in Type 1 AA9 proteins. The loops were found to result in steric congestion that affects how the Cu2+ ion interacts with cellulose. As a result, Cu2+ binding to cellulose was observed for Type 1 and not Type 2 and 3 AA9 proteins. In this study force field parameters have been evaluated for the Type 1 active site of AA9 proteins and this parameters were evaluated on all three types and binding. Future work will focus on identifying the nature of the reactive oxygen species and performing QM/MM calculations to elucidate the reactive mechanism of all three AA9 LPMO types.
- Full Text:
- Date Issued: 2018
- Authors: Moses, Vuyani
- Date: 2018
- Subjects: Copper proteins , Cellulose , Molecular dynamics , Cellulose -- Biodegradation , Bioinformatics
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/58327 , vital:27230
- Description: AA9 proteins are metallo-enzymes which are crucial for the early stages of cellulose degradation. AA9 proteins have been suggested to cleave glycosidic bonds linking cellulose through the use of their Cu2+ coordinating active site. AA9 proteins possess different regioselectivities depending on the resulting cleavage they form and as result, are grouped accordingly. Type 1 AA9 proteins cleave the C1 carbon of cellulose while Type 2 AA9 proteins cleave the C4 carbon and Type 3 AA9 proteins cleave either C1 or C4 carbons. The steric congestion of the AA9 active site has been proposed to be a contributor to the observed regioselectivity. As such, a bioinformatics characterisation of type-specific sequence and structural features was performed. Initially AA9 protein sequences were obtained from the Pfam database and multiple sequence alignment was performed. The sequences were phylogenetically characterised and sequences were grouped into their respective types and sub-groups were identified. A selection analysis was performed on AA9 LPMO types to determine the selective pressure acting on AA9 protein residues. Motif discovery was then performed to identify conserved sequence motifs in AA9 proteins. Once type-specific sequence features were identified structural mapping was performed to assess possible effects on substrate interaction. Physicochemical property analysis was also performed to assess biochemical differences between AA9 LPMO types. Molecular dynamics (MD) simulations were then employed to dynamically assess the consequences of the discovered type-specific features on AA9-cellulose interaction. Due to the absence of AA9 specific force field parameters MD simulations were not readily applicable. As a result, Potential Energy Surface (PES) scans were performed to evaluate the force field parameters for the AA9 active site using the PM6 semi empirical approach and least squares fitting. A Type 1 AA9 active site was constructed from the crystal structure 4B5Q, encompassing only the Cu2+ coordinating residues, the Cu2+ ion and two water residues. Due to the similarity in AA9 active sites, the Type force field parameters were validated on all three AA9 LPMO types. Two MD simulations for each AA9 LPMO types were conducted using two separate Lennard-Jones parameter sets. Once completed, the MD trajectories were analysed for various features including the RMSD, RMSF, radius of gyration, coordination during simulation, hydrogen bonding, secondary structure conservation and overall protein movement. Force field parameters were successfully evaluated and validated for AA9 proteins. MD simulations of AA9 proteins were able to reveal the presence of unique type-specific binding modes of AA9 active sites to cellulose. These binding modes were characterised by the presence of unique type-specific loops which were present in Type 2 and 3 AA9 proteins but not in Type 1 AA9 proteins. The loops were found to result in steric congestion that affects how the Cu2+ ion interacts with cellulose. As a result, Cu2+ binding to cellulose was observed for Type 1 and not Type 2 and 3 AA9 proteins. In this study force field parameters have been evaluated for the Type 1 active site of AA9 proteins and this parameters were evaluated on all three types and binding. Future work will focus on identifying the nature of the reactive oxygen species and performing QM/MM calculations to elucidate the reactive mechanism of all three AA9 LPMO types.
- Full Text:
- Date Issued: 2018
Synthesis and biolgical screening of potential plasmodium falciparum DXR inhibitors
- Authors: Adeyemi, Christiana Modupe
- Date: 2017-04
- Subjects: Plasmodium falciparum , Enzyme inhibitors , Malaria , Antimalarials , Drug development , Malaria -- Chemotherapy , Isopentenoids -- Synthesis , Fosmidomycin , 1-Deoxy-D-xylulose 5-phosphate
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/61790 , vital:28060
- Description: The non-mevalonate isoprenoid pathway, also known as the 1-deoxy-D-xylulose-5- phosphate DXP pathway, is absent in humans, but present in the anopheles mosquito responsible for the transmission of malaria. DXP reductoisomerase - a key enzyme in the DXP pathway in Plasmodium falciparum (PfDXR) has been identified as a target for the design of novel anti-malarial drugs. Fosmidomycin and its acetyl analogue (FR900098) are known to be inhibitors of PfDXR and, in this study, synthetic variations of the fosmidomycin scaffold have led to four series of novel analogues. Particular attention has been centred on the introduction of various substituted benzyl groups in each of these series in order to occupy a recently discovered vacant pocket in the PfDXR active-site and thus enhance ligand-enzyme binding. In the process 160 ligands and precursors have been prepared, no less than 119 of them novel. Fistly, a series of C-benzylated phosphonate esters and phosphonic acids were synthesised, in which the fosmidomycin hydroxamate Mg2+- coordinating moiety was replaced by an amide funtionality and the number of methylene groups in the “hydrophobic patch” between the phosphonate and the hydroxamate moiety was decreased from two to one. Several approaches were explored for this series, the most successful involving reaction of 3- substituted anilines with a-bromo propanoic acid in the presence of the coupling agent 1,1'- carbonyldiimidazole (CDI), followed by Michaelis-Arbuzov phosphonation using triethyl phosphite. Reaction of the resulting chiral phosphonate esters with bromotrimethylsilane gave the corresponding phosphonic acids in good yields. In order to obviate chirality issues, a second series of potential “reverse” fosmidomycin analogues was synthesised by replacing the methylene group adjacent to the the phosphonate moiety with a nitrogen atom. Deprotonation, alkylation and phosphorylation of various amines gave diethyl #-benzylphosphoramidate ester intermediate. Aza-Michael addition of these intermediates, followed by hydrolysis gave the corresponding carboxylic acids which could be reacted with different hydroxylamine hydrochloride derivatives to afford the novel hydroxamic acid derivatives in good yields. Thirdly, a series of a novel #-benzylated phosphoramidate derivatives were prepared as aza- FR900098 analogues. Alkylation of different amines using bromoacetalde-hyde diethylacetal gave a series of N-benzyl-2,2-diethoxyethylamine compounds, which were then elaborated via a futher six steps to afford novel #-benzylated phosphoramidate derivatives. Finally, in order to ensure syn-orientation of the donor atoms in the Mg - coordinating group and, at the same time, introduce conformational constraints in the ligand, the hydrophobic patch and the hydroxamate moiety were replaced by cyclic systems. Several approaches towards the synthesis of such conformationally constrained phosphoramidate analogues from maleic anhydride led to the unexpected isolation of an unprecedented acyclic furfuryl compound, and 1H NMR and DFT-level theoretical studies have been initiated to explore the reaction sequence. A series of #-benzylated phosphoramidate derivatives containing dihydroxy aromatic rings (as the conformationally constrained groups) to replace the hydroxamate moiety, were successfully obtained in six steps from the starting material, 3,4-dihydroxylbenzaldehyde. While in vitro assays have been conducted on all of the synthesised compounds, and some of the ligands show promising anti-malarial inhibitory activity - most especially the conformationally constrained cyclic #-benzylated phosphoramidate series. Interestingly, a number of these compounds has also shown activity against T.brucei - the causative agent of sleeping sickness. In silico docking studies of selected compounds has revealed the capacity of some of the ligands to bind effectively in the PfDXR active-site with the newly introduced benzyl group occupying the adjacent vacant pocket. The physico-chemical properties of these ligands were also explored in order to predict the oral-bioavailability. Most of the ligands obeyed the Lipinski rule of 5, while QSAR methods have been used in an attempt to correlate structural variations and calculated molecular properties with the bioassay data. , Thesis (PhD) -- Faculty of Science, Chemistry, 2017
- Full Text:
- Date Issued: 2017-04
- Authors: Adeyemi, Christiana Modupe
- Date: 2017-04
- Subjects: Plasmodium falciparum , Enzyme inhibitors , Malaria , Antimalarials , Drug development , Malaria -- Chemotherapy , Isopentenoids -- Synthesis , Fosmidomycin , 1-Deoxy-D-xylulose 5-phosphate
- Language: English
- Type: Doctoral theses , text
- Identifier: http://hdl.handle.net/10962/61790 , vital:28060
- Description: The non-mevalonate isoprenoid pathway, also known as the 1-deoxy-D-xylulose-5- phosphate DXP pathway, is absent in humans, but present in the anopheles mosquito responsible for the transmission of malaria. DXP reductoisomerase - a key enzyme in the DXP pathway in Plasmodium falciparum (PfDXR) has been identified as a target for the design of novel anti-malarial drugs. Fosmidomycin and its acetyl analogue (FR900098) are known to be inhibitors of PfDXR and, in this study, synthetic variations of the fosmidomycin scaffold have led to four series of novel analogues. Particular attention has been centred on the introduction of various substituted benzyl groups in each of these series in order to occupy a recently discovered vacant pocket in the PfDXR active-site and thus enhance ligand-enzyme binding. In the process 160 ligands and precursors have been prepared, no less than 119 of them novel. Fistly, a series of C-benzylated phosphonate esters and phosphonic acids were synthesised, in which the fosmidomycin hydroxamate Mg2+- coordinating moiety was replaced by an amide funtionality and the number of methylene groups in the “hydrophobic patch” between the phosphonate and the hydroxamate moiety was decreased from two to one. Several approaches were explored for this series, the most successful involving reaction of 3- substituted anilines with a-bromo propanoic acid in the presence of the coupling agent 1,1'- carbonyldiimidazole (CDI), followed by Michaelis-Arbuzov phosphonation using triethyl phosphite. Reaction of the resulting chiral phosphonate esters with bromotrimethylsilane gave the corresponding phosphonic acids in good yields. In order to obviate chirality issues, a second series of potential “reverse” fosmidomycin analogues was synthesised by replacing the methylene group adjacent to the the phosphonate moiety with a nitrogen atom. Deprotonation, alkylation and phosphorylation of various amines gave diethyl #-benzylphosphoramidate ester intermediate. Aza-Michael addition of these intermediates, followed by hydrolysis gave the corresponding carboxylic acids which could be reacted with different hydroxylamine hydrochloride derivatives to afford the novel hydroxamic acid derivatives in good yields. Thirdly, a series of a novel #-benzylated phosphoramidate derivatives were prepared as aza- FR900098 analogues. Alkylation of different amines using bromoacetalde-hyde diethylacetal gave a series of N-benzyl-2,2-diethoxyethylamine compounds, which were then elaborated via a futher six steps to afford novel #-benzylated phosphoramidate derivatives. Finally, in order to ensure syn-orientation of the donor atoms in the Mg - coordinating group and, at the same time, introduce conformational constraints in the ligand, the hydrophobic patch and the hydroxamate moiety were replaced by cyclic systems. Several approaches towards the synthesis of such conformationally constrained phosphoramidate analogues from maleic anhydride led to the unexpected isolation of an unprecedented acyclic furfuryl compound, and 1H NMR and DFT-level theoretical studies have been initiated to explore the reaction sequence. A series of #-benzylated phosphoramidate derivatives containing dihydroxy aromatic rings (as the conformationally constrained groups) to replace the hydroxamate moiety, were successfully obtained in six steps from the starting material, 3,4-dihydroxylbenzaldehyde. While in vitro assays have been conducted on all of the synthesised compounds, and some of the ligands show promising anti-malarial inhibitory activity - most especially the conformationally constrained cyclic #-benzylated phosphoramidate series. Interestingly, a number of these compounds has also shown activity against T.brucei - the causative agent of sleeping sickness. In silico docking studies of selected compounds has revealed the capacity of some of the ligands to bind effectively in the PfDXR active-site with the newly introduced benzyl group occupying the adjacent vacant pocket. The physico-chemical properties of these ligands were also explored in order to predict the oral-bioavailability. Most of the ligands obeyed the Lipinski rule of 5, while QSAR methods have been used in an attempt to correlate structural variations and calculated molecular properties with the bioassay data. , Thesis (PhD) -- Faculty of Science, Chemistry, 2017
- Full Text:
- Date Issued: 2017-04
Generation of a virtual library of terpenes using graph theory, and its application in exploration of the mechanisms of terpene biosynthesis
- Authors: Dendera, Washington
- Date: 2020
- Subjects: Terpenes , Plants -- Metabolism , Computational biology , Bioinformatics , Organic compounds -- Synthesis , Monoterpenes , Molecular biology -- Computer simulation
- Language: English
- Type: text , Thesis , Masters , MSc
- Identifier: http://hdl.handle.net/10962/123453 , vital:35439
- Description: Terpenes form a large group of organic compounds which have proven to be of use to many living organisms being used by plants for metabolism (Pichersky and Gershenzon, 1934; McGarvey and Croteau, 1995; Gershenzon and Dudareva, 2007), defence or as a means to attract pollinators and also used by humans in medical, pharmaceutical and food industry (Bicas, Dionísio and Pastore, 2009; Marmulla and Harder, 2014; Kandi et al., 2015). Following on literature methods to generate chemical libraries using graph theoretic techniques, complete libraries of all possible terpene isomers have been constructed with the goal of construction of derivative libraries of possible carbocation intermediates which are important in the elucidation of mechanisms in the biosynthesis of terpenes. Virtual library generation of monoterpenes was first achieved by generating graphs of order 7, 8, 9 and 10 using the Nauty and Traces suite. These were screened and processed with a set of collated Python scripts written to recognize the graphs in text format and translate them to molecules, minimizing through Tinker whilst discarding graphs that violate chemistry laws. As a result of the computational time required only order 7 and order 10 graphs were processed. Out of the 873 graphs generated from order seven, 353 were converted to molecules and from the 11,7 million produced from order 10 half were processed resulting in the production of 442928 compounds (repeats included). For screening, 55 366 compounds were docked in the active site of limonene synthase; of these 2355 ligands had a good Vina docking score with a binding energy of between -7.0 and -7.4 kcal.mol-1. When these best docked molecules were overlaid in the active site a map of possible ligand positions within the active site of limonene synthase was traced out.
- Full Text:
- Date Issued: 2020
- Authors: Dendera, Washington
- Date: 2020
- Subjects: Terpenes , Plants -- Metabolism , Computational biology , Bioinformatics , Organic compounds -- Synthesis , Monoterpenes , Molecular biology -- Computer simulation
- Language: English
- Type: text , Thesis , Masters , MSc
- Identifier: http://hdl.handle.net/10962/123453 , vital:35439
- Description: Terpenes form a large group of organic compounds which have proven to be of use to many living organisms being used by plants for metabolism (Pichersky and Gershenzon, 1934; McGarvey and Croteau, 1995; Gershenzon and Dudareva, 2007), defence or as a means to attract pollinators and also used by humans in medical, pharmaceutical and food industry (Bicas, Dionísio and Pastore, 2009; Marmulla and Harder, 2014; Kandi et al., 2015). Following on literature methods to generate chemical libraries using graph theoretic techniques, complete libraries of all possible terpene isomers have been constructed with the goal of construction of derivative libraries of possible carbocation intermediates which are important in the elucidation of mechanisms in the biosynthesis of terpenes. Virtual library generation of monoterpenes was first achieved by generating graphs of order 7, 8, 9 and 10 using the Nauty and Traces suite. These were screened and processed with a set of collated Python scripts written to recognize the graphs in text format and translate them to molecules, minimizing through Tinker whilst discarding graphs that violate chemistry laws. As a result of the computational time required only order 7 and order 10 graphs were processed. Out of the 873 graphs generated from order seven, 353 were converted to molecules and from the 11,7 million produced from order 10 half were processed resulting in the production of 442928 compounds (repeats included). For screening, 55 366 compounds were docked in the active site of limonene synthase; of these 2355 ligands had a good Vina docking score with a binding energy of between -7.0 and -7.4 kcal.mol-1. When these best docked molecules were overlaid in the active site a map of possible ligand positions within the active site of limonene synthase was traced out.
- Full Text:
- Date Issued: 2020
An in-silico investigation of Morita-Baylis-Hillman accessible heterocyclic analogues for applications as novel HIV-1 C protease inhibitors
- Authors: Sigauke, Lester Takunda
- Date: 2015
- Subjects: Protease inhibitors , Heterocyclic compounds , HIV (Viruses) , HIV infections , Drug resistance , Cheminformatics
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4152 , http://hdl.handle.net/10962/d1017913
- Description: Cheminformatic approaches have been employed to optimize the bis-coumarin scaffold identified by Onywera et al. (2012) as a potential hit against the protease HIV-1 protein. The Open Babel library of commands was used to access functions that were incorporated into a markov chain recursive program that generated 17750 analogues of the bis-coumarin scaffold. The Morita-Baylis-Hillman accessible heterocycles were used to introduce structural diversity within the virtual library. In silico high through-put virtual screening using AutoDock Vina was used to rapidly screen the virtual library ligand set against 61 protease models built by Onywera et al. (2012). CheS-Mapper computed a principle component analysis of the compounds based on 13 selected chemical descriptors. The compounds were plotted against the principle component analysis within a 3 dimensional chemical space in order to inspect the diversity of the virtual library. The physicochemical properties and binding affinities were used to identify the top 3 performing ligands. ACPYPE was used to inspect the constitutional properties and eliminated virtual compounds that possessed open valences. Chromene based ligand 805 and ligand 6610 were selected as the lead candidates from the high-throughput virtual screening procedure we employed. Molecular dynamic simulations of the lead candidates performed for 5 ns allowed the stability of the ligand protein complexes with protease model 305152. The free energy of binding of the leads with protease model 305152 was computed over the first 50 ps of simulation using the molecular mechanics Poisson-Boltzmann method. Analysis structural features and energy profiles from molecular dynamic simulations of the protein–ligand complexes indicated that although ligand 805 had a weaker binding affinity in terms of docking, it outperformed ligand 6610 in terms of complex stability and free energy of binding. Medicinal chemistry approaches will be used to optimize the lead candidates before their analogues will be synthesized and assayed for in vivo protease activity.
- Full Text:
- Date Issued: 2015
- Authors: Sigauke, Lester Takunda
- Date: 2015
- Subjects: Protease inhibitors , Heterocyclic compounds , HIV (Viruses) , HIV infections , Drug resistance , Cheminformatics
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:4152 , http://hdl.handle.net/10962/d1017913
- Description: Cheminformatic approaches have been employed to optimize the bis-coumarin scaffold identified by Onywera et al. (2012) as a potential hit against the protease HIV-1 protein. The Open Babel library of commands was used to access functions that were incorporated into a markov chain recursive program that generated 17750 analogues of the bis-coumarin scaffold. The Morita-Baylis-Hillman accessible heterocycles were used to introduce structural diversity within the virtual library. In silico high through-put virtual screening using AutoDock Vina was used to rapidly screen the virtual library ligand set against 61 protease models built by Onywera et al. (2012). CheS-Mapper computed a principle component analysis of the compounds based on 13 selected chemical descriptors. The compounds were plotted against the principle component analysis within a 3 dimensional chemical space in order to inspect the diversity of the virtual library. The physicochemical properties and binding affinities were used to identify the top 3 performing ligands. ACPYPE was used to inspect the constitutional properties and eliminated virtual compounds that possessed open valences. Chromene based ligand 805 and ligand 6610 were selected as the lead candidates from the high-throughput virtual screening procedure we employed. Molecular dynamic simulations of the lead candidates performed for 5 ns allowed the stability of the ligand protein complexes with protease model 305152. The free energy of binding of the leads with protease model 305152 was computed over the first 50 ps of simulation using the molecular mechanics Poisson-Boltzmann method. Analysis structural features and energy profiles from molecular dynamic simulations of the protein–ligand complexes indicated that although ligand 805 had a weaker binding affinity in terms of docking, it outperformed ligand 6610 in terms of complex stability and free energy of binding. Medicinal chemistry approaches will be used to optimize the lead candidates before their analogues will be synthesized and assayed for in vivo protease activity.
- Full Text:
- Date Issued: 2015
Synthesis and bioassay of rationally designed DXR inhibitors as potential antimalarial lead compounds
- Authors: Nokalipa, Iviwe Cwaita
- Date: 2017
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: http://hdl.handle.net/10962/4888 , vital:20740
- Description: Globally, the eradication of malaria has been challenging due to the problem of resistance that past and currently available drugs exhibit. This is exacerbated by the inherent need for anti-malarial drugs to be affordable to the poverty-stricken majority that is primarily affected by this burden. This research has focused on the development of potential inhibitors of 1-deoxy-D- xylulose-5 phosphate reductoisomerase (DXR), an essential enzyme in the mevalonate- independent pathway for the biosynthesis of isoprenoids in Plasmodium falciparum. DXR mediates the isomerisation and reduction of 1-deoxy-D-xylulose-5-phosphate into 2-C- methyl-D-erithrytol 4-phosphate. This enzyme has been determined to be a target for the development of novel antimalarial agents and extensive molecular modelling has been undertaken to develop inhibitors that fit into the DXR active site. The in silico docking data have been used to inform the design and synthesis of various N-benzyl-substituted phosphoramidate ligands that were determined to have potential as novel substrate mimics of fosmidomycin, a known DXR inhibitor. Synthesis of the N-benzyl-substituted phosphoramidate ligands involved a nine-step sequence commencing from diethyl phosphoramidate. In all, some 40 compounds have been prepared, some of them new, and were fully characterized using NMR. Attention has also been given to the mass spectrometric fragmentation patterns exhibited by selected intermediates. Four of the final products were evaluated for in vitro antimalarial activity using a PLDH assay and exhibited IC50 values < 100 µM.
- Full Text:
- Date Issued: 2017
- Authors: Nokalipa, Iviwe Cwaita
- Date: 2017
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: http://hdl.handle.net/10962/4888 , vital:20740
- Description: Globally, the eradication of malaria has been challenging due to the problem of resistance that past and currently available drugs exhibit. This is exacerbated by the inherent need for anti-malarial drugs to be affordable to the poverty-stricken majority that is primarily affected by this burden. This research has focused on the development of potential inhibitors of 1-deoxy-D- xylulose-5 phosphate reductoisomerase (DXR), an essential enzyme in the mevalonate- independent pathway for the biosynthesis of isoprenoids in Plasmodium falciparum. DXR mediates the isomerisation and reduction of 1-deoxy-D-xylulose-5-phosphate into 2-C- methyl-D-erithrytol 4-phosphate. This enzyme has been determined to be a target for the development of novel antimalarial agents and extensive molecular modelling has been undertaken to develop inhibitors that fit into the DXR active site. The in silico docking data have been used to inform the design and synthesis of various N-benzyl-substituted phosphoramidate ligands that were determined to have potential as novel substrate mimics of fosmidomycin, a known DXR inhibitor. Synthesis of the N-benzyl-substituted phosphoramidate ligands involved a nine-step sequence commencing from diethyl phosphoramidate. In all, some 40 compounds have been prepared, some of them new, and were fully characterized using NMR. Attention has also been given to the mass spectrometric fragmentation patterns exhibited by selected intermediates. Four of the final products were evaluated for in vitro antimalarial activity using a PLDH assay and exhibited IC50 values < 100 µM.
- Full Text:
- Date Issued: 2017
Mechanistic studies of unusual Miruta-Baylis-Hillman reactions
- Authors: Nyoni, Dubekile
- Date: 2012
- Subjects: Chemical reactions Benzaldehyde Acrylonitrile Methyl acrylate Coumarins
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4400 , http://hdl.handle.net/10962/d1006692
- Description: This study has focussed on the application of synthetic, kinetic and exploratory theoretical methods in elucidating the reaction mechanisms of four Morita-Baylis-Hillman (MBH) type reactions, viz, i) the reactions of the disulfide 2,2'-dithiodibenzaldehyde with various activated alkenes in the presence of DBU and Ph₃P, ii) the reactions of chromone-3-carbaldehydes with MVK, iii) the reactions of chromone-2-carbaldehydes with acrylonitrile and iv) with methyl acrylate. Attention has also been given to the origin of the observed regioselectivity in Michaelis-Arbuzov reactions of 3-(halomethyl)coumarins. Cleavage of the sulfur-sulfur bond of aryl and heteroaryl disulfides by the nitrogen nucleophile DBU has been demonstrated, and a dramatic increase in the rate of tandem MBH and disulfide cleavage reactions of 2,2'-dithiodibenzaldehyde with the activated alkenes, MVK, EVK, acrylonitrile, methyl acrylate and tert-butyl acrylate has been achieved through the use of the dual organo-catalyst system, DBU-Ph₃P – an improvement accompanied by an increase in the yields of the isolated products. Detailed NMR-based kinetic studies have been conducted on the DBU-catalysed reactions of 2,2'-dithiodibenzaldehyde with MVK and methyl acrylate, and a theoretical kinetic model has been developed and complementary computational studies using Gaussian 03, at the DFT-B3LYP/6-31G(d) level of theory have provided valuable insights into the mechanism of these complex transformations. Reactions of chromone-3-carbaldehydes with MVK to afford chromone dimers and tricyclic products have been repeated, and a novel, intermediate MBH adduct has been isolated. The mechanisms of the competing pathways have been elucidated by DFT calculations and the development of a detailed theoretical kinetic model has ensued. Unusual transformations in MBH-type reactions of chromone-2-carbaldehydes with acrylonitrile and methyl acrylate have been explored and the structures of the unexpected products have been established using 1- and 2-D NMR, HRMS and X-ray crystallographic techniques. Attention has also been given to the synthesis of 3-(halomethyl)coumarins via the MBH reaction, and their subsequent Michaelis-Arbuzov reactions with triethyl phosphite. An exploratory study of the kinetics of the phosphonation reaction has been undertaken and the regio-selectivity of nucleophilic attack at the 4- and 1'-positions in the 3-chloro- and 3-(iodomethyl)coumarins has been investigated by calculating Mulliken charges, LUMO surfaces and Fukui condensed local softness functions.
- Full Text:
- Date Issued: 2012
- Authors: Nyoni, Dubekile
- Date: 2012
- Subjects: Chemical reactions Benzaldehyde Acrylonitrile Methyl acrylate Coumarins
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4400 , http://hdl.handle.net/10962/d1006692
- Description: This study has focussed on the application of synthetic, kinetic and exploratory theoretical methods in elucidating the reaction mechanisms of four Morita-Baylis-Hillman (MBH) type reactions, viz, i) the reactions of the disulfide 2,2'-dithiodibenzaldehyde with various activated alkenes in the presence of DBU and Ph₃P, ii) the reactions of chromone-3-carbaldehydes with MVK, iii) the reactions of chromone-2-carbaldehydes with acrylonitrile and iv) with methyl acrylate. Attention has also been given to the origin of the observed regioselectivity in Michaelis-Arbuzov reactions of 3-(halomethyl)coumarins. Cleavage of the sulfur-sulfur bond of aryl and heteroaryl disulfides by the nitrogen nucleophile DBU has been demonstrated, and a dramatic increase in the rate of tandem MBH and disulfide cleavage reactions of 2,2'-dithiodibenzaldehyde with the activated alkenes, MVK, EVK, acrylonitrile, methyl acrylate and tert-butyl acrylate has been achieved through the use of the dual organo-catalyst system, DBU-Ph₃P – an improvement accompanied by an increase in the yields of the isolated products. Detailed NMR-based kinetic studies have been conducted on the DBU-catalysed reactions of 2,2'-dithiodibenzaldehyde with MVK and methyl acrylate, and a theoretical kinetic model has been developed and complementary computational studies using Gaussian 03, at the DFT-B3LYP/6-31G(d) level of theory have provided valuable insights into the mechanism of these complex transformations. Reactions of chromone-3-carbaldehydes with MVK to afford chromone dimers and tricyclic products have been repeated, and a novel, intermediate MBH adduct has been isolated. The mechanisms of the competing pathways have been elucidated by DFT calculations and the development of a detailed theoretical kinetic model has ensued. Unusual transformations in MBH-type reactions of chromone-2-carbaldehydes with acrylonitrile and methyl acrylate have been explored and the structures of the unexpected products have been established using 1- and 2-D NMR, HRMS and X-ray crystallographic techniques. Attention has also been given to the synthesis of 3-(halomethyl)coumarins via the MBH reaction, and their subsequent Michaelis-Arbuzov reactions with triethyl phosphite. An exploratory study of the kinetics of the phosphonation reaction has been undertaken and the regio-selectivity of nucleophilic attack at the 4- and 1'-positions in the 3-chloro- and 3-(iodomethyl)coumarins has been investigated by calculating Mulliken charges, LUMO surfaces and Fukui condensed local softness functions.
- Full Text:
- Date Issued: 2012
Enumeration, conformation sampling and population of libraries of peptide macrocycles for the search of chemotherapeutic cardioprotection agents
- Authors: Sigauke, Lester Takunda
- Date: 2019
- Subjects: Peptides -- Synthesis , Macrocyclic compounds , Drug development , Drug discovery , Cardiovascular system -- Diseases -- Prevention , Proteins -- Synthesis
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/116056 , vital:34293
- Description: Peptides are uniquely endowed with features that allow them to perturb previously difficult to drug biomolecular targets. Peptide macrocycles in particular have seen a flurry of recent interest due to their enhanced bioavailability, tunability and specificity. Although these properties make them attractive hit-candidates in early stage drug discovery, knowing which peptides to pursue is non‐trivial due to the magnitude of the peptide sequence space. Computational screening approaches show promise in their ability to address the size of this search space but suffer from their inability to accurately interrogate the conformational landscape of peptide macrocycles. We developed an in‐silico compound enumerator that was tasked with populating a conformationally laden peptide virtual library. This library was then used in the search for cardio‐protective agents (that may be administered, reducing tissue damage during reperfusion after ischemia (heart attacks)). Our enumerator successfully generated a library of 15.2 billion compounds, requiring the use of compression algorithms, conformational sampling protocols and management of aggregated compute resources in the context of a local cluster. In the absence of experimental biophysical data, we performed biased sampling during alchemical molecular dynamics simulations in order to observe cyclophilin‐D perturbation by cyclosporine A and its mitochondrial targeted analogue. Reliable intermediate state averaging through a WHAM analysis of the biased dynamic pulling simulations confirmed that the cardio‐protective activity of cyclosporine A was due to its mitochondrial targeting. Paralleltempered solution molecular dynamics in combination with efficient clustering isolated the essential dynamics of a cyclic peptide scaffold. The rapid enumeration of skeletons from these essential dynamics gave rise to a conformation laden virtual library of all the 15.2 Billion unique cyclic peptides (given the limits on peptide sequence imposed). Analysis of this library showed the exact extent of physicochemical properties covered, relative to the bare scaffold precursor. Molecular docking of a subset of the virtual library against cyclophilin‐D showed significant improvements in affinity to the target (relative to cyclosporine A). The conformation laden virtual library, accessed by our methodology, provided derivatives that were able to make many interactions per peptide with the cyclophilin‐D target. Machine learning methods showed promise in the training of Support Vector Machines for synthetic feasibility prediction for this library. The synergy between enumeration and conformational sampling greatly improves the performance of this library during virtual screening, even when only a subset is used.
- Full Text:
- Date Issued: 2019
- Authors: Sigauke, Lester Takunda
- Date: 2019
- Subjects: Peptides -- Synthesis , Macrocyclic compounds , Drug development , Drug discovery , Cardiovascular system -- Diseases -- Prevention , Proteins -- Synthesis
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/116056 , vital:34293
- Description: Peptides are uniquely endowed with features that allow them to perturb previously difficult to drug biomolecular targets. Peptide macrocycles in particular have seen a flurry of recent interest due to their enhanced bioavailability, tunability and specificity. Although these properties make them attractive hit-candidates in early stage drug discovery, knowing which peptides to pursue is non‐trivial due to the magnitude of the peptide sequence space. Computational screening approaches show promise in their ability to address the size of this search space but suffer from their inability to accurately interrogate the conformational landscape of peptide macrocycles. We developed an in‐silico compound enumerator that was tasked with populating a conformationally laden peptide virtual library. This library was then used in the search for cardio‐protective agents (that may be administered, reducing tissue damage during reperfusion after ischemia (heart attacks)). Our enumerator successfully generated a library of 15.2 billion compounds, requiring the use of compression algorithms, conformational sampling protocols and management of aggregated compute resources in the context of a local cluster. In the absence of experimental biophysical data, we performed biased sampling during alchemical molecular dynamics simulations in order to observe cyclophilin‐D perturbation by cyclosporine A and its mitochondrial targeted analogue. Reliable intermediate state averaging through a WHAM analysis of the biased dynamic pulling simulations confirmed that the cardio‐protective activity of cyclosporine A was due to its mitochondrial targeting. Paralleltempered solution molecular dynamics in combination with efficient clustering isolated the essential dynamics of a cyclic peptide scaffold. The rapid enumeration of skeletons from these essential dynamics gave rise to a conformation laden virtual library of all the 15.2 Billion unique cyclic peptides (given the limits on peptide sequence imposed). Analysis of this library showed the exact extent of physicochemical properties covered, relative to the bare scaffold precursor. Molecular docking of a subset of the virtual library against cyclophilin‐D showed significant improvements in affinity to the target (relative to cyclosporine A). The conformation laden virtual library, accessed by our methodology, provided derivatives that were able to make many interactions per peptide with the cyclophilin‐D target. Machine learning methods showed promise in the training of Support Vector Machines for synthetic feasibility prediction for this library. The synergy between enumeration and conformational sampling greatly improves the performance of this library during virtual screening, even when only a subset is used.
- Full Text:
- Date Issued: 2019
Large and multi scale mechanistic modeling of Diels-Alder reactions
- Authors: Isamura, Bienfait Kabuyaya
- Date: 2022-04-06
- Subjects: Computational chemistry , Diels-Alder reaction , Python (Computer program language) , Reaction force theory , Fullerenes , Diolefins , AMADAR (Automated workflow for Mechanistic Analysis of Diels-Alder Reactions , ONIOM
- Language: English
- Type: Master's thesis , text
- Identifier: http://hdl.handle.net/10962/232317 , vital:49981
- Description: The [4+2] cycloaddition reaction between conjugated dienes and substituted alkenes is known as the Diels-Alder (DA) reaction, in honor of two German chemists, Otto Diels and Kurt Alder, who first reported this marvelous chemical transformation. The DA reaction is one of the most popular reactions in organic chemistry, allowing for the regio- and stereospecific establishment of six-membered rings with up to four stereogenic centers. This pericyclic reaction has found many applications in areas as diverse as natural products chemistry, polymer chemistry, and agrochemistry. Over the past decades, the mechanism of the Diels-Alder (DA) reaction has been the subject of numerous studies, dealing with questions as diverse as the mechanistic pathway, the synchronicity, the use of catalysts, the effect of solvents and salts, etc. On the other hand, as an example, fullerenes (and particularly [60] fullerene) have been found to act as good dienophiles in DA reactions to the extent that many functionalized fullerenes with interesting applications are still synthesized by reacting C60 with dienes. However, despite the very abundant literature about the mechanism of the DA reaction, some pertinent questions have been still pending, including, without being restricted to, the prediction of transition state (TS) geometries and the modeling of DA reactions involving large systems, such as those of C60 fullerene. It must be emphasized that TSs are not easy to predict and the main reason is that many existing algorithms require that the search is initiated from a good starting point (guess TS), which must be very similar to the actual TS. This problem is even more difficult when many TSs are to be located as may be the case in large-scale studies. Moreover, due to the large size of the C60 molecule, the usage of accurate high-level computational methods in the investigation of its reactivity towards dienes is computationally costly, implying the need to find the best threshold between accuracy and computational cost. Therefore, the present study was carried out to contribute to solving the problems of large-scale prediction of DA transition state geometries and the multi-scale modeling of C60 fullerene DA reactions. To address the first problem (large-scale prediction of TSs), we have developed a python program named “AMADAR”, which predicts an unlimited number of DA transition states, using only the SMILES strings of the cycloadducts. AMADAR is customizable and allows for the description of intramolecular DA reactions as well as systems resulting in competing paths. In addition, The AMADAR tool contains two separate modules that perform reaction force analyses and atomic decomposition of energy derivatives from the predicted Intrinsic Reaction Coordinates (IRC) paths. The performance of AMADAR was assessed using 2000 DA cycloadducts and showed a success rate of ~ 95%. Most of the errors were due to basis set inconsistencies or convergence issues that we are still working on. Furthermore, a set of 150 IRC paths generated by the AMADAR program were analyzed to get insight into the (a)synchronicity of DA reactions. This investigation confirmed that the reaction force constant 𝜅 (second derivatives of the system energy with respect to the reaction coordinate) was a good indicator of synchronicity in DA reactions. A close inspection of the profile of 𝜅 has enabled us to propose an alternative classification of DA reactions based on their synchronicity degree, in terms of (quasi)-synchronous, moderate asynchronous, asynchronous, and likely two-steps DA reactions. Natural population analyses seemed to indicate that the global maximum of the reaction force constant could be identified with the formation of all the bonds in the reaction site. Finally, the atomic resolution of energy derivatives suggested that the mechanism of the DA reaction involves two inner elementary processes associated with the formation of each C-C bond. A striking mechanistic difference between synchronous and asynchronous DA reactions emerging from this study is that, in asynchronous reactions, the driving and retarding forces are mainly caused by the fast and slow-forming bonds (elementary process) respectively, while in the case of synchronous ones both elementary processes retard and drive the process concomitantly and equivalently. Regarding the DA reaction of C60 fullerene that was considered to illustrate the problem of multiscale modeling, we have constructed 12 ONIOM2 and 10 ONIOM3 models combining five semi-empirical methods (AM1, PM3, PM3MM, PDDG, PM6) and the LDA(SVWN) functional in conjunction with the B3LYP/6-31G(d) level. Then, their accuracy and efficiency were assessed in comparison with the pure B3LYP/6-31G(d) level considering first the DA reaction between C60 and cyclopentadiene whose experimental data are available. Further, different DFT functionals were employed in place of the B3LYP functional to describe the higher-layer of the best ONIOM partition, and the results obtained were compared to experimental data. At this step, the ONIOM2(M06-2X/6-31 G(d): SVWN/STO-3G) model, where the higher layer encompasses the diene and pyracyclene portion of C60, was found to provide the best tradeoff between accuracy and cost, with respect to experimental data. This model showed errors lower than 2.6 and 2.0 kcal/mol for the estimation of the activation and reaction enthalpies respectively. We have also demonstrated, by comparing several ONIOM2(DFT/6-31G(d): SVWN/STO-3G) models, the importance of dispersion corrections in the accurate estimation of reaction and activation energies. Finally, we have considered a set of 21 dienes, including anthracene, 1,3-butadiene, 1,3-cyclopentadiene, furan, thiophene, selenothiophene, pyrrole and their mono-cyano and hydroxyl derivatives to get insight into the DA reaction of C60 using the best ONIOM2(M06-2X/6-31 G(d): SVWN/STO-3G) model. For a given diene and its derivatives, the analysis of frontier molecular orbitals provides a consistent explanation for the substituent effect on the activation barrier. It revealed that electron-donating (withdrawing) groups such as -OH (–CN) cut down on the activation barrier of the reaction by lowering (extending) of the HOMOdiene – LUMOC60 gap and consequently enhancing (weakening) the interaction between the two reactants. Further, the decomposition of the activation energy into the strain and interaction components suggested that, for a given diene, electron-donating groups (here –OH) diminish the height of the activation barrier not only by favoring the attractive interaction between the diene and C60, but also by reducing the strain energy of the system; the opposite effect is observed for electron-withdrawing groups (here –CN). In contrast with some previous findings on typical DA reactions, we could not infer any general rule applicable to the entire dataset for the prediction of activation energies because the latter does not correlate well with either of the TS polarity, electrophilicity of the diene, or the reaction energy. , Thesis (MSc) -- Faculty of Science, Chemistry, 2022
- Full Text:
- Date Issued: 2022-04-06
- Authors: Isamura, Bienfait Kabuyaya
- Date: 2022-04-06
- Subjects: Computational chemistry , Diels-Alder reaction , Python (Computer program language) , Reaction force theory , Fullerenes , Diolefins , AMADAR (Automated workflow for Mechanistic Analysis of Diels-Alder Reactions , ONIOM
- Language: English
- Type: Master's thesis , text
- Identifier: http://hdl.handle.net/10962/232317 , vital:49981
- Description: The [4+2] cycloaddition reaction between conjugated dienes and substituted alkenes is known as the Diels-Alder (DA) reaction, in honor of two German chemists, Otto Diels and Kurt Alder, who first reported this marvelous chemical transformation. The DA reaction is one of the most popular reactions in organic chemistry, allowing for the regio- and stereospecific establishment of six-membered rings with up to four stereogenic centers. This pericyclic reaction has found many applications in areas as diverse as natural products chemistry, polymer chemistry, and agrochemistry. Over the past decades, the mechanism of the Diels-Alder (DA) reaction has been the subject of numerous studies, dealing with questions as diverse as the mechanistic pathway, the synchronicity, the use of catalysts, the effect of solvents and salts, etc. On the other hand, as an example, fullerenes (and particularly [60] fullerene) have been found to act as good dienophiles in DA reactions to the extent that many functionalized fullerenes with interesting applications are still synthesized by reacting C60 with dienes. However, despite the very abundant literature about the mechanism of the DA reaction, some pertinent questions have been still pending, including, without being restricted to, the prediction of transition state (TS) geometries and the modeling of DA reactions involving large systems, such as those of C60 fullerene. It must be emphasized that TSs are not easy to predict and the main reason is that many existing algorithms require that the search is initiated from a good starting point (guess TS), which must be very similar to the actual TS. This problem is even more difficult when many TSs are to be located as may be the case in large-scale studies. Moreover, due to the large size of the C60 molecule, the usage of accurate high-level computational methods in the investigation of its reactivity towards dienes is computationally costly, implying the need to find the best threshold between accuracy and computational cost. Therefore, the present study was carried out to contribute to solving the problems of large-scale prediction of DA transition state geometries and the multi-scale modeling of C60 fullerene DA reactions. To address the first problem (large-scale prediction of TSs), we have developed a python program named “AMADAR”, which predicts an unlimited number of DA transition states, using only the SMILES strings of the cycloadducts. AMADAR is customizable and allows for the description of intramolecular DA reactions as well as systems resulting in competing paths. In addition, The AMADAR tool contains two separate modules that perform reaction force analyses and atomic decomposition of energy derivatives from the predicted Intrinsic Reaction Coordinates (IRC) paths. The performance of AMADAR was assessed using 2000 DA cycloadducts and showed a success rate of ~ 95%. Most of the errors were due to basis set inconsistencies or convergence issues that we are still working on. Furthermore, a set of 150 IRC paths generated by the AMADAR program were analyzed to get insight into the (a)synchronicity of DA reactions. This investigation confirmed that the reaction force constant 𝜅 (second derivatives of the system energy with respect to the reaction coordinate) was a good indicator of synchronicity in DA reactions. A close inspection of the profile of 𝜅 has enabled us to propose an alternative classification of DA reactions based on their synchronicity degree, in terms of (quasi)-synchronous, moderate asynchronous, asynchronous, and likely two-steps DA reactions. Natural population analyses seemed to indicate that the global maximum of the reaction force constant could be identified with the formation of all the bonds in the reaction site. Finally, the atomic resolution of energy derivatives suggested that the mechanism of the DA reaction involves two inner elementary processes associated with the formation of each C-C bond. A striking mechanistic difference between synchronous and asynchronous DA reactions emerging from this study is that, in asynchronous reactions, the driving and retarding forces are mainly caused by the fast and slow-forming bonds (elementary process) respectively, while in the case of synchronous ones both elementary processes retard and drive the process concomitantly and equivalently. Regarding the DA reaction of C60 fullerene that was considered to illustrate the problem of multiscale modeling, we have constructed 12 ONIOM2 and 10 ONIOM3 models combining five semi-empirical methods (AM1, PM3, PM3MM, PDDG, PM6) and the LDA(SVWN) functional in conjunction with the B3LYP/6-31G(d) level. Then, their accuracy and efficiency were assessed in comparison with the pure B3LYP/6-31G(d) level considering first the DA reaction between C60 and cyclopentadiene whose experimental data are available. Further, different DFT functionals were employed in place of the B3LYP functional to describe the higher-layer of the best ONIOM partition, and the results obtained were compared to experimental data. At this step, the ONIOM2(M06-2X/6-31 G(d): SVWN/STO-3G) model, where the higher layer encompasses the diene and pyracyclene portion of C60, was found to provide the best tradeoff between accuracy and cost, with respect to experimental data. This model showed errors lower than 2.6 and 2.0 kcal/mol for the estimation of the activation and reaction enthalpies respectively. We have also demonstrated, by comparing several ONIOM2(DFT/6-31G(d): SVWN/STO-3G) models, the importance of dispersion corrections in the accurate estimation of reaction and activation energies. Finally, we have considered a set of 21 dienes, including anthracene, 1,3-butadiene, 1,3-cyclopentadiene, furan, thiophene, selenothiophene, pyrrole and their mono-cyano and hydroxyl derivatives to get insight into the DA reaction of C60 using the best ONIOM2(M06-2X/6-31 G(d): SVWN/STO-3G) model. For a given diene and its derivatives, the analysis of frontier molecular orbitals provides a consistent explanation for the substituent effect on the activation barrier. It revealed that electron-donating (withdrawing) groups such as -OH (–CN) cut down on the activation barrier of the reaction by lowering (extending) of the HOMOdiene – LUMOC60 gap and consequently enhancing (weakening) the interaction between the two reactants. Further, the decomposition of the activation energy into the strain and interaction components suggested that, for a given diene, electron-donating groups (here –OH) diminish the height of the activation barrier not only by favoring the attractive interaction between the diene and C60, but also by reducing the strain energy of the system; the opposite effect is observed for electron-withdrawing groups (here –CN). In contrast with some previous findings on typical DA reactions, we could not infer any general rule applicable to the entire dataset for the prediction of activation energies because the latter does not correlate well with either of the TS polarity, electrophilicity of the diene, or the reaction energy. , Thesis (MSc) -- Faculty of Science, Chemistry, 2022
- Full Text:
- Date Issued: 2022-04-06
Novel applications of Morita-Baylis-Hillman methodology in organic synthesis
- Authors: Mciteka, Lulama Patrick
- Date: 2013 , 2013-04-22
- Subjects: Organic compounds -- Synthesis -- Research Asymmetric synthesis Asymmetry (Chemistry) Chemical reactions -- Research Camphor -- Research AZT (Drug) -- Research Chemical inhibitors -- Research Chemistry -- Methodology
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4439 , http://hdl.handle.net/10962/d1007598
- Description: The overall approach in the present investigation has been to explore applications of the Morita-Baylis-Hillman (MBH) reaction in asymmetric synthesis and in the continuation of systems with medicinal potential. To this end, a series of varied camphor-derived acrylate esters was prepared to serve as chiral substrates in asymmetric Morita-Baylis- Hillman reactions. Reduction of N-substituted camphor-10-sulfonamides afforded the 3- exo-hydroxy derivatives as the major products. Acylation of the corresponding sodium alkoxides gave the desired 3-exo-acrylate esters, isolation of which was complicated by concomitant formation of hydrochlorinated and diastereomeric competition products. Bulky camphorsulfonamides containing alkyl, dialkyl, aromatic and adamantyl groups were selected as N-substituents with the view of achieving stereoselective outcome in subsequent MBH reactions. The synthesis of novel camphor-derived Morita-Baylis-Hillman adducts using various pyridine-carboxaldehydes proceeded with exceptionally high yields with diastereoselectivities ranging from 7-33 % d.e. Both 1D and 2D NMR and HRMS techniques were employed to confirm the structures and an extensive study of the electropositive fragmentation patterns of a number of camphor-derived chiral acrylate esters was conducted. Attention has also been given to the application of MBH methodology in the construction of heterocyclic ‘cinnamate-like’ AZT conjugates which were designed to serve as dualaction HIV-1 integrase-reverse transcriptase (IN-RT) inhibitors. A number of pyridine carboxaldehyde-derived MBH adducts were synthesized using methyl, ethyl and t-butyl acrylates in the presence of 3-hydroxyquinuclidine (3-HQ) as catalyst. The yields for these reactions were excellent. The resulting MBH adducts were acetylated and subjected to aza-Michael addition using propargylamine. The resulting alkylamino compounds were then used in ‘Click reactions’ to form the targeted AZT-conjugates in moderate to excellent yield. In silico docking of computer modelled AZT-conjugates into the HIV-1 integrase and reverse transcriptase enzyme-active sites and potential hydrogen-bonding interaction with active-site amino acid residues were identified. The electrospray MS fragmentations of the AZT and the novel AZT-conjugates were also investigated and common fragmentation pathways were identified.
- Full Text:
- Date Issued: 2013
- Authors: Mciteka, Lulama Patrick
- Date: 2013 , 2013-04-22
- Subjects: Organic compounds -- Synthesis -- Research Asymmetric synthesis Asymmetry (Chemistry) Chemical reactions -- Research Camphor -- Research AZT (Drug) -- Research Chemical inhibitors -- Research Chemistry -- Methodology
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4439 , http://hdl.handle.net/10962/d1007598
- Description: The overall approach in the present investigation has been to explore applications of the Morita-Baylis-Hillman (MBH) reaction in asymmetric synthesis and in the continuation of systems with medicinal potential. To this end, a series of varied camphor-derived acrylate esters was prepared to serve as chiral substrates in asymmetric Morita-Baylis- Hillman reactions. Reduction of N-substituted camphor-10-sulfonamides afforded the 3- exo-hydroxy derivatives as the major products. Acylation of the corresponding sodium alkoxides gave the desired 3-exo-acrylate esters, isolation of which was complicated by concomitant formation of hydrochlorinated and diastereomeric competition products. Bulky camphorsulfonamides containing alkyl, dialkyl, aromatic and adamantyl groups were selected as N-substituents with the view of achieving stereoselective outcome in subsequent MBH reactions. The synthesis of novel camphor-derived Morita-Baylis-Hillman adducts using various pyridine-carboxaldehydes proceeded with exceptionally high yields with diastereoselectivities ranging from 7-33 % d.e. Both 1D and 2D NMR and HRMS techniques were employed to confirm the structures and an extensive study of the electropositive fragmentation patterns of a number of camphor-derived chiral acrylate esters was conducted. Attention has also been given to the application of MBH methodology in the construction of heterocyclic ‘cinnamate-like’ AZT conjugates which were designed to serve as dualaction HIV-1 integrase-reverse transcriptase (IN-RT) inhibitors. A number of pyridine carboxaldehyde-derived MBH adducts were synthesized using methyl, ethyl and t-butyl acrylates in the presence of 3-hydroxyquinuclidine (3-HQ) as catalyst. The yields for these reactions were excellent. The resulting MBH adducts were acetylated and subjected to aza-Michael addition using propargylamine. The resulting alkylamino compounds were then used in ‘Click reactions’ to form the targeted AZT-conjugates in moderate to excellent yield. In silico docking of computer modelled AZT-conjugates into the HIV-1 integrase and reverse transcriptase enzyme-active sites and potential hydrogen-bonding interaction with active-site amino acid residues were identified. The electrospray MS fragmentations of the AZT and the novel AZT-conjugates were also investigated and common fragmentation pathways were identified.
- Full Text:
- Date Issued: 2013
Activity of diverse chalcones against several targets: statistical analysis of a high-throughput virtual screen of a custom chalcone library
- Authors: Sarron, Arthur F D
- Date: 2020
- Subjects: Acetophenone , Benzaldehyde , Ketones , Pyruvate kinase , Drug development , Aromatic compounds , Heat shock proteins
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/116028 , vital:34291
- Description: Chalcone family molecules are well known to have therapeutic proprieties (anti-inflammatory, anti-microbial or anti-cancer, etc). However the mechanism of action in some cases is not well known. A virtual library of this family of compounds was constructed using custom scripts, based on the aldol condensation, and this library was modified further to analogues by expansion of the α,β-unsaturated ketone linker. Acetophenone and benzaldehyde derivatives which are available and purchasable were used as a base to design the chalcone virtual library. 8063 chalcones were constructed and geometrically optimized with Gaussian 09. Their physicochemical characteristics linked to the Lipinski rules were analyzed with Knime and CDK. The entire library was after docked against several targets including HIV-1 integrase, MRSA pyruvate kinase, HSP90, COX-1, COX-2, ALR2, MAOA, MAOB, acetylcholinesterase, butyrylcholinesterase and PLA2. With the exception of MAOA, which does not have a crystal structure ligand, all dockings were validated by redocking the original ligand provided by the literature. These targets are known in the literature to be inhibited by chalcone-derivatives. However, specificity of the particular known chalcone inhibitors to the particular targets is not known. To this end the performance of the generated chalcone library against the list of targets was of interest. The binding energy of ligand-protein complexes was generally good across the library. Statistical analysis including principal component analysis and hierarchical clustering analysis were made in order to investigate for any physical/chemical characteristics which might explain what chalcone features affect the binding energy of the ligand-protein complexes. The spherical polar coordinates defining the orientation of the binding poses were also calculated and used in the statistical analysis. The statistical analysis has allowed us to hypothesize the importance of these radial distances and the polar angles of key atoms in the chalcones in binding to the pyruvate kinase crystal structure. This was validated by the docking of another small library of compound models in which the α,β-unsaturated ketone chain of the chalcone was replaced by incrementally longer conjugated chains. Further studies on the chalcones themselves reveal rotameric systems in both cis and trans-configurations (which may impact binding), and also studied was the effect of Topliss-based modification and its impact of binding to HSP90. Molecular dynamics confirmed good binding of identified chalcone hits.
- Full Text:
- Date Issued: 2020
- Authors: Sarron, Arthur F D
- Date: 2020
- Subjects: Acetophenone , Benzaldehyde , Ketones , Pyruvate kinase , Drug development , Aromatic compounds , Heat shock proteins
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/116028 , vital:34291
- Description: Chalcone family molecules are well known to have therapeutic proprieties (anti-inflammatory, anti-microbial or anti-cancer, etc). However the mechanism of action in some cases is not well known. A virtual library of this family of compounds was constructed using custom scripts, based on the aldol condensation, and this library was modified further to analogues by expansion of the α,β-unsaturated ketone linker. Acetophenone and benzaldehyde derivatives which are available and purchasable were used as a base to design the chalcone virtual library. 8063 chalcones were constructed and geometrically optimized with Gaussian 09. Their physicochemical characteristics linked to the Lipinski rules were analyzed with Knime and CDK. The entire library was after docked against several targets including HIV-1 integrase, MRSA pyruvate kinase, HSP90, COX-1, COX-2, ALR2, MAOA, MAOB, acetylcholinesterase, butyrylcholinesterase and PLA2. With the exception of MAOA, which does not have a crystal structure ligand, all dockings were validated by redocking the original ligand provided by the literature. These targets are known in the literature to be inhibited by chalcone-derivatives. However, specificity of the particular known chalcone inhibitors to the particular targets is not known. To this end the performance of the generated chalcone library against the list of targets was of interest. The binding energy of ligand-protein complexes was generally good across the library. Statistical analysis including principal component analysis and hierarchical clustering analysis were made in order to investigate for any physical/chemical characteristics which might explain what chalcone features affect the binding energy of the ligand-protein complexes. The spherical polar coordinates defining the orientation of the binding poses were also calculated and used in the statistical analysis. The statistical analysis has allowed us to hypothesize the importance of these radial distances and the polar angles of key atoms in the chalcones in binding to the pyruvate kinase crystal structure. This was validated by the docking of another small library of compound models in which the α,β-unsaturated ketone chain of the chalcone was replaced by incrementally longer conjugated chains. Further studies on the chalcones themselves reveal rotameric systems in both cis and trans-configurations (which may impact binding), and also studied was the effect of Topliss-based modification and its impact of binding to HSP90. Molecular dynamics confirmed good binding of identified chalcone hits.
- Full Text:
- Date Issued: 2020
Nanocomposites of nickel phthalocyanines as electrocatalysts for the oxidation of chlorophenols an experimental and theoretical approach
- Authors: Khene, Mielie Samson
- Date: 2012
- Subjects: Nanocomposites (Materials) -- Research Phthalocyanines -- Research Chlorophenols -- Research Electrocatalysis -- Research
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4326 , http://hdl.handle.net/10962/d1004986
- Description: In this work the interaction between peripherally (b) substituted nickel tetrahydroxyphthalocyanines (b-NiPc(OH)4 and poly-b-Ni(O)Pc(OH)4) with 4-chlorophenol is theoretically rationalized by performing calculations at the B3LYP/6-31G(d) level. Density functional theory (DFT) and molecular orbital theory are used to calculate the condensed Fukui function for phthalocyanine derivatives and 4-chlorophenol, in order to determine the reactive sites involved when 4-chlorophenol is oxidized, and to compare theoretically predicted reactivity to experimentally determined electrocatalytic activity. Electrocatalytic activities of adsorbed NiPc derivatives: OPGE-α-NiPc(OH)8 (OPGE = ordinary pyrolytic graphite electrode, α = nonperipheral substitution), OPGE-a-NiPc(OH)4 and OPGE-β- NiPc(OH)4 are compared with those of the polymerized counterparts: OPGE-poly-α-Ni(O)Pc(OH)8, OPGE poly-a-NiPc(OH)4 and OPGE-poly-β-NiPc(OH)4, respectively. β-NiPc(NH2)4, β-NiPc(OH)4, α-NiPc(OH)4, α-NiPc(OH)8, α-NiPc(C10H21)8 are adsorbed on singled walled carbon nanotube (SWCNT) and β-NiPc(NH2)4-SWCNT (linked to SWCNT), are used to modify glassy carbon electrode (GCE) and employed for the electro oxidation of chlorophenols. The β-NiPc(NH2)4-SWCNT gave the best current response for the oxidation of 4-chlorophenol. α-NiPc(OH)8-SWCNT and α-NiPc(C10H21)8 gave the best resistance to electrode fouling due to oxidation by product of 4-chlorophenol. The synthesis of conjugates of cadmium telluride quantum dots (CdTe-QDs) capped with thioglycolic acid and peripherally substituted nickel tetraamino phthalocyanine (β-NiPc(NH2)4) complex were also employed for the oxidation of chlorophenols. Separation of one of the diastereomers of 1, 2-subnaphthalocyanine (SubNPc) was achieved as well as separation of the enantiomers. The absorption and magnetic circular dichroism spectra, together with theoretical calculations, reveal rather small variations in the frontier molecular orbitals of the SubNPc compared to conventional subphthalocyanine (SubPc), except for the destabilization of the highest occupied molecular orbital (HOMO), which results in a characteristic absorption in the Soret band region. The chirality of SubNPc, including the cyclic dichroism (CD) signs and intensities, are discussed in detail.
- Full Text:
- Date Issued: 2012
- Authors: Khene, Mielie Samson
- Date: 2012
- Subjects: Nanocomposites (Materials) -- Research Phthalocyanines -- Research Chlorophenols -- Research Electrocatalysis -- Research
- Language: English
- Type: Thesis , Doctoral , PhD
- Identifier: vital:4326 , http://hdl.handle.net/10962/d1004986
- Description: In this work the interaction between peripherally (b) substituted nickel tetrahydroxyphthalocyanines (b-NiPc(OH)4 and poly-b-Ni(O)Pc(OH)4) with 4-chlorophenol is theoretically rationalized by performing calculations at the B3LYP/6-31G(d) level. Density functional theory (DFT) and molecular orbital theory are used to calculate the condensed Fukui function for phthalocyanine derivatives and 4-chlorophenol, in order to determine the reactive sites involved when 4-chlorophenol is oxidized, and to compare theoretically predicted reactivity to experimentally determined electrocatalytic activity. Electrocatalytic activities of adsorbed NiPc derivatives: OPGE-α-NiPc(OH)8 (OPGE = ordinary pyrolytic graphite electrode, α = nonperipheral substitution), OPGE-a-NiPc(OH)4 and OPGE-β- NiPc(OH)4 are compared with those of the polymerized counterparts: OPGE-poly-α-Ni(O)Pc(OH)8, OPGE poly-a-NiPc(OH)4 and OPGE-poly-β-NiPc(OH)4, respectively. β-NiPc(NH2)4, β-NiPc(OH)4, α-NiPc(OH)4, α-NiPc(OH)8, α-NiPc(C10H21)8 are adsorbed on singled walled carbon nanotube (SWCNT) and β-NiPc(NH2)4-SWCNT (linked to SWCNT), are used to modify glassy carbon electrode (GCE) and employed for the electro oxidation of chlorophenols. The β-NiPc(NH2)4-SWCNT gave the best current response for the oxidation of 4-chlorophenol. α-NiPc(OH)8-SWCNT and α-NiPc(C10H21)8 gave the best resistance to electrode fouling due to oxidation by product of 4-chlorophenol. The synthesis of conjugates of cadmium telluride quantum dots (CdTe-QDs) capped with thioglycolic acid and peripherally substituted nickel tetraamino phthalocyanine (β-NiPc(NH2)4) complex were also employed for the oxidation of chlorophenols. Separation of one of the diastereomers of 1, 2-subnaphthalocyanine (SubNPc) was achieved as well as separation of the enantiomers. The absorption and magnetic circular dichroism spectra, together with theoretical calculations, reveal rather small variations in the frontier molecular orbitals of the SubNPc compared to conventional subphthalocyanine (SubPc), except for the destabilization of the highest occupied molecular orbital (HOMO), which results in a characteristic absorption in the Soret band region. The chirality of SubNPc, including the cyclic dichroism (CD) signs and intensities, are discussed in detail.
- Full Text:
- Date Issued: 2012