Ruben Cloete
University of the Western Cape, SANBI, Graduate Student
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Research Interests: Computer Science, Computational Biology, Biology, Drug Discovery, Medicine, and 10 moreBiological Sciences, Tuberculosis, Humans, Mycobacterium tuberculosis, Mathematical Sciences, Drug Resistance, BMC Bioinformatics, Quantitative Structure Activity Relationship, Anti-Bacterial Agents, and Metabolic pathway
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Research Interests: Engineering, Genetics, Chemistry, Comparative sequence analysis, Molecular Evolution, and 15 moreBiology, Medicine, Multidisciplinary, High Frequency, Humans, Mycobacterium tuberculosis, Comparative Analysis, Gene, Mycobacterium tuberculosis complex, Genetic variation, Genome sequence, Gene Family, Cell Surface Markers, Cohort Studies, and Antigenic Variation
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HIV-1 is responsible for a spectrum of neurocognitive deficits defined as HIV-associated neurocognitive disorders (HAND). The HIV transactivator of transcription (Tat) protein plays a key role in the neuropathophysiology of HAND. The Tat... more
HIV-1 is responsible for a spectrum of neurocognitive deficits defined as HIV-associated neurocognitive disorders (HAND). The HIV transactivator of transcription (Tat) protein plays a key role in the neuropathophysiology of HAND. The Tat protein functions by transactivation of viral genes through its interaction with the transactivation response (TAR) RNA element. Subtype-specific Tat protein signatures including C31S, R57S and Q63E present in Tat subtype C has previously been linked to a lowered neuropathophysiology compared to Tat subtype B. In this study, we attempted to understand the molecular mechanism by which Tat subtype-specific variation, particularly, C31S, R57S, and Q63E influence the Tat-TAR interaction. We performed molecular modeling to generate accurate three-dimensional protein structures of the HIV-1 Tat subtypes C and B using the Swiss model webserver. Thereafter, we performed a molecular docking of the TAR RNA element to each of the Tat subtypes B and C protein s...
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Parkinson’s disease (PD), the fastest-growing neurological disorder globally, has a complex etiology. A previous study by our group identified the p.G849D variant in neurexin 2 (NRXN2), encoding the synaptic protein, NRXN2α, as a possible... more
Parkinson’s disease (PD), the fastest-growing neurological disorder globally, has a complex etiology. A previous study by our group identified the p.G849D variant in neurexin 2 (NRXN2), encoding the synaptic protein, NRXN2α, as a possible causal variant of PD. Therefore, we aimed to perform functional studies using proteomics in an attempt to understand the biological pathways affected by the variant. We hypothesized that this may reveal insight into the pathobiology of PD. Wild-type and mutant NRXN2α plasmids were transfected into SH-SY5Y cells. Thereafter, total protein was extracted and prepared for mass spectrometry using a Thermo Scientific Fusion mass spectrometer equipped with a Nanospray Flex ionization source. The data were then interrogated against the UniProt H. sapiens database and afterward, pathway and enrichment analyses were performed using in silico tools. Overexpression of the wild-type protein led to the enrichment of proteins involved in neurodegenerative disease...
Research Interests: Biology and Proteomics
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Synaptopathies are brain disorders characterized by dysfunctional synapses, which are specialized junctions between neurons and essential for the transmission of information. Synaptic dysfunction can occur due to mutations that alter the... more
Synaptopathies are brain disorders characterized by dysfunctional synapses, which are specialized junctions between neurons and essential for the transmission of information. Synaptic dysfunction can occur due to mutations that alter the structure and function of synaptic components or abnormal expression levels of a synaptic protein. One class of synaptic proteins that are essential to their biology are cell adhesion proteins that connect the pre- and post-synaptic compartments. Neurexins are one type of synaptic cell adhesion molecule that have, recently, gained more pathological interest. Variants in both neurexins and their common binding partners, neuroligins, have been associated with several neuropsychiatric disorders. In this review, we summarized some of the key physiological functions of the neurexin protein family and the protein networks they are involved in. Furthermore, examination of published literature has implicated neurexins in both neuropsychiatric and neurodegen...
GO terms for interacting partners of neurexins. A) Biological processes. B) Cellular components. C) Molecular functions. All: total number of proteins analyzed. The number above each bar indicates the number of proteins assigned to that... more
GO terms for interacting partners of neurexins. A) Biological processes. B) Cellular components. C) Molecular functions. All: total number of proteins analyzed. The number above each bar indicates the number of proteins assigned to that category. Figure generated by WebGestalt (http://www.webgestalt.org) [18];GeneMANIA Report;Physiological and pathway data for interacting partners of neurexins
Docking scores and number of interactions for the top ten compounds to Rv1712. The residues highlighted in bold are conserved catalytic residues while ligand C5P is highlighted in red. The numbers inside the () indicate the number of... more
Docking scores and number of interactions for the top ten compounds to Rv1712. The residues highlighted in bold are conserved catalytic residues while ligand C5P is highlighted in red. The numbers inside the () indicate the number of interactions formed between residue and the compound atom. Data can be viewed in Microsoft excel. (XLS 8 kb)
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KEGG pathway descriptors for M. tuberculosis genes retained after prioritization. Data can be viewed in Microsoft excel. (XLS 7 kb)
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Blast hits obtained for Rv1712, Rv2984, Rv2194, Rv1311, Rv1305, Rv2195, Rv1622c, Rv1456c and Rv2421c against three host intestinal bacteria. Bacterial species highlighted in bold showed homology to the query gene. Data can be viewed in... more
Blast hits obtained for Rv1712, Rv2984, Rv2194, Rv1311, Rv1305, Rv2195, Rv1622c, Rv1456c and Rv2421c against three host intestinal bacteria. Bacterial species highlighted in bold showed homology to the query gene. Data can be viewed in Microsoft excel. (XLS 13 kb)
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The 24 sequenced Mycobacterium strains. Data can be viewed in Microsoft excel. (XLS 34 kb)
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The variation in total (green) and potential energy (red) for the Rv1712-C5P complex during the 30000Â ps simulation. Generated using Gnuplotv4.2 [41]. (PDF 69 kb)
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KEGG metabolic pathway map for Oxidative phosphorylation in M. tuberculosis H37rV strain. Rv2984 or ppk selected for investigation is shown in blue highlighted box and involved in step 2.7.4.1 of this specific pathway. Rv2194 or qcrC... more
KEGG metabolic pathway map for Oxidative phosphorylation in M. tuberculosis H37rV strain. Rv2984 or ppk selected for investigation is shown in blue highlighted box and involved in step 2.7.4.1 of this specific pathway. Rv2194 or qcrC selected for investigation is shown in the red highlighted box. Both Rv1305 (atpE) and Rv1311 (atpC) selected for investigation are shown in brown highlighted box and involved in step 3.6.3.14 of this specific pathway. Rv2195 or qcrA selected for investigation is shown in the yellow highlighted box involved in cytochrome C reductase. Rv1456c or COX15 selected for investigation is shown in the magenta highlighted box involved in cytochrome C oxidase. Rv1622c or CydB selected for investigation is shown in the light blue highlighted box involved in cytochrome C oxidase. Known drug resistance gene Rv1854c or ndh is shown in orange highlighted box and involved in step 1.6.99.3 of this pathway. M. tuberculosis specific genes are coloured in green. (PDF 157 kb)
Research Interests: Pharmacology, Biochemistry, Genetics, Marine Biology, Microbiology, and 15 moreNeuroscience, Physiology, Immunology, Molecular Biology, Computational Biology, Plant Biology, Cancer, Cell Biology, Infectious Diseases, Investigation, Cytochrome c oxidase, COX, Oxidative phosphorylation, Rv, and Drug target identification
RMSD of the backbone atoms of model Rv1712 (green) and substrate C5P (red) during the 30000Â ps simulation. Generated using Gnuplotv4.2 [41]. (PDF 198 kb)
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KEGG pathways for genes known to be associated with 1st and 2nd line drug resistance. The 10 mutations (eight genes) indicated in bold were verified by the BROAD institute, while the remainder were identified from literature.... more
KEGG pathways for genes known to be associated with 1st and 2nd line drug resistance. The 10 mutations (eight genes) indicated in bold were verified by the BROAD institute, while the remainder were identified from literature. Abbreviations used: A-Adenine, Ala-Alanine, Arg-Arginine, Asn-Glutamine, Asp-Aspartate, Gly-Glycine, His-Histidine, indel-insertion deletion, Ileu-Isoleucine, Leu-Leucine, Met-Methionine, N.A-Not available, Pro-Proline, Ser-Serine, T-Thymidine, Thr-Threonine, Tyr-Tyrosine, Val-Valine. (PDF 143 kb)
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Radius of gyration of all bacbone atoms for Rv1712 over the 30000Â ps simulation. (PDF 29 kb)
Nicotinamide-nucleotide adenylyl transferase (Rv2421c) was selected as a potential drug target, because it has been shown, in vitro, to be essential for Mycobacterium tuberculosis growth. It is conserved between mycobacterium species, is... more
Nicotinamide-nucleotide adenylyl transferase (Rv2421c) was selected as a potential drug target, because it has been shown, in vitro, to be essential for Mycobacterium tuberculosis growth. It is conserved between mycobacterium species, is up-regulated during dormancy, has a known 3D crystal structure and has no known human homologs. A model of Rv2421c in complex with nicotinic acid adenine dinucleotide and magnesium ion was constructed and subject to virtual ligand screening against the Prestwick Chemical Library and the ZINC database, which yielded 155 potential hit molecules. 3D-QSAR studies of the 155 drug molecules indicated five compounds with similar inhibitory efficiencies compared to known inhibitors of Rv2421c. Molecular docking validation and molecular dynamics simulation analysis of the top five compounds indicated that the identified inhibitor molecules bind to Rv2421c with comparable efficiency as the substrate DND. Subsequent in vitro testing of the five compounds ident...
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Nicotinamide-nucleotide adenylyl transferase (Rv2421c) was selected as a potential drug target, because it has been shown, in vitro, to be essential for Mycobacterium tuberculosis growth. It is conserved between mycobacterium species, is... more
Nicotinamide-nucleotide adenylyl transferase (Rv2421c) was selected as a potential drug target, because it has been shown, in vitro, to be essential for Mycobacterium tuberculosis growth. It is conserved between mycobacterium species, is up-regulated during dormancy, has a known 3D crystal structure and has no known human homologs. A model of Rv2421c in complex with nicotinic acid adenine dinucleotide and magnesium ion was constructed and subject tovirtual ligand screening against the Prestwick Chemical Library and the ZINC database, which yielded 155 potential hit molecules. Of the 155 compounds identified five were pursued further using an IC50 based 3D-QSAR study. The 3D-QSAR model validated the inhibition properties of the five compounds based on R2 value of 0.895 and Q2 value of 0.944 compared to known inhibitors of Rv2421c. Higher binding affinities was observed for the novel ZINC13544129 and two FDA approved compounds (Novobiocin sodium salt, Sulfasalazine). Similarly, the to...
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Parkinson’s disease (PD) is a neurodegenerative disorder exhibiting Mendelian inheritance in some families. Next-generation sequencing approaches, including whole exome sequencing (WES), have revolutionized the field of Mendelian... more
Parkinson’s disease (PD) is a neurodegenerative disorder exhibiting Mendelian inheritance in some families. Next-generation sequencing approaches, including whole exome sequencing (WES), have revolutionized the field of Mendelian disorders and have identified a number of PD genes. We recruited a South African family with autosomal dominant PD and used WES to identify a possible pathogenic mutation. After filtration and prioritization, we found five potential causative variants inCFAP65,RTF1,NRXN2,TEP1andCCNF. The variant inNRXN2was selected for further analysis based on consistent prediction of deleteriousness across computational tools, not being present in unaffected family members, ethnic-matched controls or public databases, and its expression in the substantia nigra. A protein model for NRNX2 was created which provided a three-dimensional (3D) structure that satisfied qualitative mean and global model quality assessment scores. Trajectory analysis showed destabilizing effects o...
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Synaptopathies are brain disorders characterized by dysfunctional synapses, which are specialized junctions between neurons that are essential for the transmission of information. Synaptic dysfunction can occur due to mutations that alter... more
Synaptopathies are brain disorders characterized by dysfunctional synapses, which are specialized junctions between neurons that are essential for the transmission of information. Synaptic dysfunction can occur due to mutations that alter the structure and function of synaptic components or abnormal expression levels of a synaptic protein. One class of synaptic proteins that are essential to their biology are cell adhesion proteins that connect the pre- and post-synaptic compartments. Neurexins are one type of synaptic cell adhesion molecule that have, recently, gained more pathological interest. Variants in both neurexins and their common binding partners, neuroligins, have been associated with several neuropsychiatric disorders. In this review, we summarize some of the key physiological functions of the neurexin protein family and the protein networks they are involved in. Furthermore, examination of published literature has implicated neurexins in both neuropsychiatric and neurod...
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Research Interests: Computer Science, Computational Biology, Biology, Drug Discovery, Medicine, and 10 moreBiological Sciences, Tuberculosis, Humans, Mycobacterium tuberculosis, Mathematical Sciences, Drug Resistance, BMC Bioinformatics, Quantitative Structure Activity Relationship, Anti-Bacterial Agents, and Metabolic pathway
Research Interests: Engineering, Genetics, Chemistry, Comparative sequence analysis, Molecular Evolution, and 15 moreBiology, Medicine, Multidisciplinary, High Frequency, Humans, Mycobacterium tuberculosis, Comparative Analysis, Gene, Mycobacterium tuberculosis complex, Genetic variation, Genome sequence, Gene Family, Cell Surface Markers, Cohort Studies, and Antigenic Variation
The less virulent human (h) coronaviruses (CoVs) 229E, NL63, OC43, and HKU1 cause mild, self-limiting respiratory tract infections, while the more virulent SARS-CoV-1, MERS-CoV, and SARS-CoV-2 have caused severe outbreaks. The CoV... more
The less virulent human (h) coronaviruses (CoVs) 229E, NL63, OC43, and HKU1 cause mild, self-limiting respiratory tract infections, while the more virulent SARS-CoV-1, MERS-CoV, and SARS-CoV-2 have caused severe outbreaks. The CoV envelope (E) protein, an important contributor to the pathogenesis of severe hCoV infections, may provide insight into this disparate severity of the disease. We, therefore, generated full-length E protein models for SARS-CoV-1 and -2, MERS-CoV, HCoV-229E, and HCoV-NL63 and docked C-terminal peptides of each model to the PDZ domain of the human PALS1 protein. The PDZ-binding motif (PBM) of the SARS-CoV-1 and -2 and MERS-CoV models adopted a more flexible, extended coil, while the HCoV-229E and HCoV-NL63 models adopted a less flexible alpha helix. All the E peptides docked to PALS1 occupied the same binding site and the more virulent hCoV E peptides generally interacted more stably with PALS1 than the less virulent ones. We hypothesize that the increased fl...
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Human (h) coronaviruses (CoVs) 229E, NL63, OC43, and HKU1 are less virulent and cause mild, self-limiting respiratory tract infections, while SARS-CoV, MERS-CoV, and SARS-CoV-2, are more virulent and have caused severe outbreaks. The CoV... more
Human (h) coronaviruses (CoVs) 229E, NL63, OC43, and HKU1 are less virulent and cause mild, self-limiting respiratory tract infections, while SARS-CoV, MERS-CoV, and SARS-CoV-2, are more virulent and have caused severe outbreaks. The CoV envelope (E) protein, an important contributor to the pathogenesis of severe hCoVs infections, may provide insight into this disparate severity of the disease. Topology prediction programs and 3D modelling software was used to predict and visualize structural aspects of the hCoV E protein related to its functions. All seven hCoV E proteins largely adopted different topologies, with some distinction between the more virulent and less virulent ones. The 3D models refined this distinction, showing the PDZ-binding motif (PBM) of SARS-CoV, MERS-CoV, and SARS-CoV-2 to be more flexible than the PBM of hCoVs 229E, NL63, OC43, and HKU1. We speculate that the increased flexibility of the PBM may provide the more virulent hCoVs with a greater degree of freedom...
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Single nucleotide polymorphisms detected in the solute carrier member family-22 has been shown to result in a variable response in the treatment of type 2 diabetes mellitus with Metformin. This study predicted a three-dimensional protein... more
Single nucleotide polymorphisms detected in the solute carrier member family-22 has been shown to result in a variable response in the treatment of type 2 diabetes mellitus with Metformin. This study predicted a three-dimensional protein structure for the SLC22A2 protein sequence using AlphaFold 2 and modelled five haplotypes within SLC22A2 protein structure observed in the Xhosa population of South Africa. The protein models were used to determine the effect(s) of haplotype variations on the transport function of Metformin and 10 other drugs by the SLC22A2 protein. Molecular dynamic simulation studies, molecular docking and interaction analysis of the five SLC22A2 haplotypes were performed in complex with the ligand 5RE in a POPC lipid bilayer to understand the mechanism of drug binding. Weakest binding free energy was found between 5RE and haplotype 1. Molecular docking studies indicated the top binding ligands as well as Metformin to bind inside the transport channel in all haplo...
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Background The Integrase (IN) strand transfer inhibitor (INSTI), Dolutegravir (DTG), has been given the green light to form part of first-line combination antiretroviral therapy (cART) by the World Health Organization (WHO). DTG... more
Background The Integrase (IN) strand transfer inhibitor (INSTI), Dolutegravir (DTG), has been given the green light to form part of first-line combination antiretroviral therapy (cART) by the World Health Organization (WHO). DTG containing regimens have shown a high genetic barrier against HIV-1 isolates carrying specific resistance mutations when compared with other class of regimens. Methods We evaluated the HIV-1 CRF02_AG IN gene sequences from Cameroon for the presence of resistance-associated mutations (RAMs) against INSTIs and naturally occurring polymorphisms (NOPs), using study sequences (n = 20) and (n = 287) sequences data derived from HIV Los Alamos National Laboratory database. The possible impact of NOPs on protein structure caused by HIV-1 CRF02_AG variations was addressed within the context of a 3D model of the HIV-1 IN complex and interaction analysis was performed using PyMol to validate DTG binding to the Wild type and seven mutant structures. Results We observed 1...
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HIV-1 Integrase (IN) is a primary target for combination antiretroviral therapy (cART). Only a limited number of studies report on the emergence of resistance-associated mutations (RAMs) in Cameroon. We observed that (1.4%) of sequence... more
HIV-1 Integrase (IN) is a primary target for combination antiretroviral therapy (cART). Only a limited number of studies report on the emergence of resistance-associated mutations (RAMs) in Cameroon. We observed that (1.4%) of sequence from treatment naïve patients had Integrase strand transfer inhibitor (INSTI) RAMs. These mutations confer resistance to raltegravir (RAL) and elvitegravir (EVG). We also observed that 10.1% of the sequences have INSTI accessory RAMs. HIV-1 CRF02_AG was the predominant subtype (44.7.%) in this study analyses. The occurrence of INSTI RAMs among the sequences at baseline needs to be monitored carefully.
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The World Health Organization (WHO) has put forth recommendations for the use of Integrase (IN) strand transfers inhibitors (INSTIs) to be part of the first-line combination antiretroviral therapy (cART) regimen to treat HIV infections.... more
The World Health Organization (WHO) has put forth recommendations for the use of Integrase (IN) strand transfers inhibitors (INSTIs) to be part of the first-line combination antiretroviral therapy (cART) regimen to treat HIV infections. The knowledge of pre-treatment drug resistance against INSTIs is still scarce in resource-limited settings. We characterised the integrase gene to identify resistance-associated mutations (RAMs) in 56 INSTI-naïve patient viral sequences from Cameroon. Study analysis used 37 sequences with fragment size ≥ 500bp or of good quality .The majority of the sequences were identified as CRF02_AG 54.% (n=20/37) and 45.9 % (n=17/37), other subtypes viral sequences includes (A, CRF36_cpx, F ,G and C). 18.9 % (n=7/37) of the sequences had RAMs, with only 5.4% (n=2/37) having major RAMs (Y143R/C/D/G and P145S), against INSTIs. Accessory RAMs were present in 8.1 % (n=3/37) of sequences, of which one sequence contained solely E157Q, another Q95K. One patient sequence had three accessory RAMs (G140E, E157Q, and G163R). We identified major RAMs to INSTIs, which might have a potential clinical impact to dolutegravir (DTG) roll out in resource-limited settings (RLS) including Cameroon. This is the first study to describe RAMs among INSTI naïve people living with HIV-1 (PLHIV-1) infected with CRF02_AG and other subtypes in Cameroon.
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Resistance associated mutations (RAMs) threaten the long-term success of combination antiretroviral therapy (cART) outcomes for HIV-1 treatment. HIV-1 Integrase (IN) strand transfer inhibitors (INSTIs) have proven to be a viable option... more
Resistance associated mutations (RAMs) threaten the long-term success of combination antiretroviral therapy (cART) outcomes for HIV-1 treatment. HIV-1 Integrase (IN) strand transfer inhibitors (INSTIs) have proven to be a viable option for highly specific HIV-1 therapy. The INSTI, Dolutegravir is recommended by the World Health Organization for use as first-line cART. This study aims to understand how RAMs affect the stability of IN, as well as the binding of the drug Dolutegravir to the catalytic pocket of the protein. Molecular modelling of HIV-1C IN was performed using the SWISS-MODEL webserver; with quality assessment performed using internal methods and external software tools. The site directed mutator webserver was used to predict destabilizing and/or stabilizing effects of known RAMs while FoldX confirmed any changes in protein energy upon introduction of mutation. Interaction analysis between neighbouring residues was done using PyMOL. Three randomly selected mutations were...
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HIV-Integrase (IN) has proven to be a viable target for highly specific HIV-1 therapy. We aimed to characterize the HIV-1 IN gene in a South African context and identify resistance-associated mutations (RAMs) against available first and... more
HIV-Integrase (IN) has proven to be a viable target for highly specific HIV-1 therapy. We aimed to characterize the HIV-1 IN gene in a South African context and identify resistance-associated mutations (RAMs) against available first and second generation Integrase strand-transfer inhibitors (InSTIs). We performed genetic analyses on 91 treatment-naïve HIV-1 infected patients, as well as 314 treatment-naive South African HIV-1 IN-sequences, downloaded from Los Alamos HIV Sequence Database. Genotypic analyses revealed the absence of major RAMs in the cohort collected before the broad availability of combination antiretroviral therapy (cART) and INSTI in South Africa, however, occurred at a rate of 2.85% (9/314) in database derived sequences. RAMs were present at IN-positions 66, 92, 143, 147 and 148, all of which may confer resistance to Raltegravir (RAL) and Elvitegravir (EVG), but are unlikely to affect second-generation Dolutegravir (DTG), except mutations in the Q148 pathway. Furt...
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A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.
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The process of viral integration into the host genome is an essential step of the HIV-1 life cycle. The viral Integrase (IN) enzyme catalyses integration. IN is an ideal therapeutic enzyme targeted by several drugs; raltegravir (RAL),... more
The process of viral integration into the host genome is an essential step of the HIV-1 life cycle. The viral Integrase (IN) enzyme catalyses integration. IN is an ideal therapeutic enzyme targeted by several drugs; raltegravir (RAL), elvitegravir (EVG), dolutegravir (DTG) and bictegravir (BIC) having been approved by the USA Food and Drug Administration (FDA). Due to high HIV-1 diversity, it is not well understood how specific naturally occurring polymorphisms (NOPs) in IN may affect the structure/function and binding affinity of Integrase Strand Transfer Inhibitors (INSTIs). In this study, we applied computational methods of molecular modelling and docking to analyse the effect of NOPs on the full-length IN structure and INSTI binding. We identified 16 NOPs within the Cameroonian derived CRF02_AG IN sequences and further identified 17 NOPs within HIV-1C South African sequences. The NOPs in the IN structures did not show any effect on INSTI binding. INSTIs displayed similar binding...