13 research outputs found
Interaction analysis of statistically enriched mutations identified in Cameroon recombinant subtype CRF02_AG that can influence the development of Dolutegravir drug resistance mutations
No full textBackground: 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 12.8% (37/287) sequences to contain RAMs, with only 1.0% (3/287) of the sequences having major INSTI RAMs: T66A, Q148H, R263K and N155H. Of these,11.8% (34/287) of the sequences contained five different IN accessory mutations; namely Q95K, T97A, G149A, E157Q and D232N. NOPs occurred at a frequency of 66% on the central core domain (CCD) position, 44% on the C-terminal domain (CTD) position and 35% of the N-terminal domain (NTD) position. The interaction analysis revealed that DTG bound to DNA, 2MG ions and DDE motif residues for T66A, T97A, Q148H, N155H and R263K comparable to the WT structure. Except for accessory mutant structure E157Q, only one MG contact was made with DTG, while DTG had no MG ion contacts and no DDE motif residue contacts for structure D232N.
Conclusions: Our analysis indicated that all RAM's that resulted in a change in the number of interactions with encompassing residues does not affect DTG binding, while accessory mutations E157Q and D232N could affect DTG binding leading to possible DTG resistance. However, further experimental validation is required to validate the in silico findings of our study.The study was supported by the National Research Foundation (NRF) of
South Africa, Poliomyelitis Research Foundation (PRF) of South Africa, Harry
Crossley Foundation, South African Research Chairs Initiative of the
Department of Science, Technology (DST). South African Medical Research
Council (SAMRC) and The Higher Education Department, next Generation of
Academic Programme (nGAP), provided support for this study in the form of
full time academic positions and salaries to R. Cloete and GBJ. Mr. Darren
Isaacs and Miss Rumbidzai Chitongo were funded by the South African
Research Chairs Initiative of the Department of Science and Innovation (DSI)
and National Research Foundation (NRF) of South Africa, award number UID
64751. The Funders had no role in the design of the study and collection,
analysis, and interpretation of data and in writing the manuscript
Investigating the structural effect of Raltegravir resistance associated mutations on the South African HIV-1 Integrase subtype C protein structure
No full text>Magister Scientiae - MScBackground and Aims Human Immunodeficiency Virus (HIV) type 1 group M subtype C (HIV-1C) accounts for nearly half of global HIV-1 infections, with South Africa (SA) being one of the countries with the highest infection burden. In recent years, SA has made great strides in tackling its HIV epidemic, resulting in the country being recognized globally as the one sub-Saharan country with the largest combination antiretroviral therapy (cART) programme. Regardless of the potency of cART, the efficacy of the treatment is limited and hampered by the emergence of drug resistance. The majority of research on HIV-1 infections, effect of antiretroviral (ARV) drugs and understanding resistance to ARV drugs has been extensively conducted, but mainly on HIV-1 subtype B (HIV-1B), with less information known about HIV-1C. HIV-1’s viral Integrase (IN) enzyme has become a viable target for highly specific cART, due to its importance in the infection and replication cycle of the virus. The lack of a complete HIV-1C IN protein structure has negatively impacted the progress on structural studies of nucleoprotein reaction intermediates. The mechanism of HIV-1 viral DNA’s integration has been studied extensively at biochemical and cellular levels, but not at a molecular level. This study aims to use in silico methods that involve molecular modeling and molecular dynamic (MD) simulations to prioritize mutations that could affect HIV-1C IN binding to DNA and the IN strand-transfer inhibitor (INSTI) dolutegravir (DTG). The purpose is to help tailor more effective personalized treatment options for patients living with HIV in SA. This study will in part use patient derived sequence data to identify mutations and model them into the protein structure to understand their impact on the HIV-1C IN protein structure folding and dynamics. Methods Our sample cohort consisted of 11 sample sequences derived from SA HIV-1 treatmentexperienced
patients who were being treated with the INSTI raltegravir (RAL). The sequences were submitted to the Stanford HIV resistance database (HIVdb) to screen for any new/novel variants resulting from possible RAL failure. Some of these new variants were analyzed to analyse their effect, if any, on the binding of DTG to the HIV-1C IN protein. Additionally, an HIV-1C IN consensus sequence constructed from SA’s HIV-1 infected population was used to model a complete three-dimensional wild type (WT) HIV-1C IN homology model. All samples were sequenced by our collaborators at the Division of Medical Virology, Stellenbosch University together with the National Health Laboratory Services (NHLS), SA. The HIV-1CZA WT-IN protein enzyme was predicted using SWISS-MODEL, and the quality of the resulting model validated. Various analyses were conducted in order to study and assess the effect of the selected new variants on the protein structure and binding of DTG to the IN protein. The mutation Cutoff Scanning Matrix (mCSM) program was used to predict protein stability after mutation, while PyMol helped to study any changes in polar contact activity before and after mutation. PyMol was also used to generate four mutant HIV-1C IN complex structures and these structures together with the WT IN were subjected to production MD simulations for 150 nanoseconds (ns). Trajectory analyses of the MD simulations were also conducted and reported. Results A total of 21 new variants were detected in our sample cohort, from which only six were chosen for further analyses within the study. A homology model of HIV-1C IN was
successfully constructed and validated. The structural quality assessment indicated high reliability of the HIV-1C IN tetrameric structure, with more than 90.0% confidence in modelled regions. Of the six selected variants, only one (S119P) was calculated to be slightly stabilizing to the protein structure, with the other five found to be destabilizing to the IN protein structure. Variant S119P showed a loss in polar contacts that could destabilize the protein structure, while variant Y143R, resulted in the gain of polar contacts which could reduce flexibility of the 140’s region affecting drug binding. Similarly, mutant systems P3 (S119P, Y143R) and P4 (V150A, M154I) showed reduced hydrogen bond formation and the weakest non-bonded pairwise interaction energy. These two systems, P3 and P4, also showed significantly reduced to none polar contacts between DTG, magnesium (MG) ions and the IN protein, compared to the WT IN and P2 mutant IN systems. Interestingly, the WT structure and systems P1 (I113V) and P2 (L63I, V75M, Y143R) showed the highest non-bonded interaction energy, compared to systems P3 and P4. This was further supported by the polar interaction analyses of simulation clusters from the WT IN and mutant IN system P2 (L63I, V75M, Y143R), which were the only protein structures that formed polar contacts with DTG, MG ions and DDE motif residues, while P1 only made contacts with DNA and IN residues. Conclusion Findings from this study leads to a conclusion that double mutants (S119P, Y143R) and (V150A, M154I) may result in a reduction in the efficacy of DTG, especially when in combination. Furthermore, variants identified in systems P1 and P2 may still allow for effective DTG binding to IN and outcompete viral DNA for host DNA to prevent strand transfer. To the best of our knowledge, this is the first study that uses the consensus WT HIV1C IN sequence to build an accurate 3D homology model to understand the effect of less frequently detected/reported variants on DTG binding in a South African context.
https://etd
Antibiotics in the Diep River and potential abatement using grape slurry waste
No full textThesis (MTech (Chemistry))--Cape Peninsula University of Technology, 2017.Pharmaceuticals have found extensive application in human health management. They are released into the environment through urine, excreta and inappropriate disposal methods. Residues of pharmaceutical products have been reported to show toxic consequences in some freshwater and marine organisms. Antibiotics are one of the most important groups of common human pharmaceuticals widely in use as prescribed and non-prescribed drugs. Antibiotics and their metabolites have been quantitated in water and found in trace levels. But even at such low concentrations they can maintain high biological activities with potential adverse effects on humans and animals. Unfortunately, many pharmaceutical compounds are resistant to breakdown in the environment, hence they have tendency for environmental magnification, since they are designed to be biologically active. Therefore, there is need to evaluate their environmental levels and their possible abatement methods using simple, cheap and low cost techniques, in order to avert their potential toxic consequences. In this research, a cost effective, robust, selective and rugged method for the analysis of antibiotics in water samples using liquid chromatography was developed, and used for monitoring levels of the selected antibiotics in Diep River. Also, an effective remediation procedure for these contaminants in water was developed using activated carbon produced from grape slurry waste
Investigating the structural effect of Raltegravir resistance associated mutations on the South African HIV-1 Integrase subtype C protein structure
No full text>Magister Scientiae - MScBackground and Aims Human Immunodeficiency Virus (HIV) type 1 group M subtype C (HIV-1C) accounts for nearly half of global HIV-1 infections, with South Africa (SA) being one of the countries with the highest infection burden. In recent years, SA has made great strides in tackling its HIV epidemic, resulting in the country being recognized globally as the one sub-Saharan country with the largest combination antiretroviral therapy (cART) programme. Regardless of the potency of cART, the efficacy of the treatment is limited and hampered by the emergence of drug resistance. The majority of research on HIV-1 infections, effect of antiretroviral (ARV) drugs and understanding resistance to ARV drugs has been extensively conducted, but mainly on HIV-1 subtype B (HIV-1B), with less information known about HIV-1C. HIV-1’s viral Integrase (IN) enzyme has become a viable target for highly specific cART, due to its importance in the infection and replication cycle of the virus. The lack of a complete HIV-1C IN protein structure has negatively impacted the progress on structural studies of nucleoprotein reaction intermediates. The mechanism of HIV-1 viral DNA’s integration has been studied extensively at biochemical and cellular levels, but not at a molecular level. This study aims to use in silico methods that involve molecular modeling and molecular dynamic (MD) simulations to prioritize mutations that could affect HIV-1C IN binding to DNA and the IN strand-transfer inhibitor (INSTI) dolutegravir (DTG). The purpose is to help tailor more effective personalized treatment options for patients living with HIV in SA. This study will in part use patient derived sequence data to identify mutations and model them into the protein structure to understand their impact on the HIV-1C IN protein structure folding and dynamics. Methods Our sample cohort consisted of 11 sample sequences derived from SA HIV-1 treatmentexperienced
patients who were being treated with the INSTI raltegravir (RAL). The sequences were submitted to the Stanford HIV resistance database (HIVdb) to screen for any new/novel variants resulting from possible RAL failure. Some of these new variants were analyzed to analyse their effect, if any, on the binding of DTG to the HIV-1C IN protein. Additionally, an HIV-1C IN consensus sequence constructed from SA’s HIV-1 infected population was used to model a complete three-dimensional wild type (WT) HIV-1C IN homology model. All samples were sequenced by our collaborators at the Division of Medical Virology, Stellenbosch University together with the National Health Laboratory Services (NHLS), SA. The HIV-1CZA WT-IN protein enzyme was predicted using SWISS-MODEL, and the quality of the resulting model validated. Various analyses were conducted in order to study and assess the effect of the selected new variants on the protein structure and binding of DTG to the IN protein. The mutation Cutoff Scanning Matrix (mCSM) program was used to predict protein stability after mutation, while PyMol helped to study any changes in polar contact activity before and after mutation. PyMol was also used to generate four mutant HIV-1C IN complex structures and these structures together with the WT IN were subjected to production MD simulations for 150 nanoseconds (ns). Trajectory analyses of the MD simulations were also conducted and reported. Results A total of 21 new variants were detected in our sample cohort, from which only six were chosen for further analyses within the study. A homology model of HIV-1C IN was
successfully constructed and validated. The structural quality assessment indicated high reliability of the HIV-1C IN tetrameric structure, with more than 90.0% confidence in modelled regions. Of the six selected variants, only one (S119P) was calculated to be slightly stabilizing to the protein structure, with the other five found to be destabilizing to the IN protein structure. Variant S119P showed a loss in polar contacts that could destabilize the protein structure, while variant Y143R, resulted in the gain of polar contacts which could reduce flexibility of the 140’s region affecting drug binding. Similarly, mutant systems P3 (S119P, Y143R) and P4 (V150A, M154I) showed reduced hydrogen bond formation and the weakest non-bonded pairwise interaction energy. These two systems, P3 and P4, also showed significantly reduced to none polar contacts between DTG, magnesium (MG) ions and the IN protein, compared to the WT IN and P2 mutant IN systems. Interestingly, the WT structure and systems P1 (I113V) and P2 (L63I, V75M, Y143R) showed the highest non-bonded interaction energy, compared to systems P3 and P4. This was further supported by the polar interaction analyses of simulation clusters from the WT IN and mutant IN system P2 (L63I, V75M, Y143R), which were the only protein structures that formed polar contacts with DTG, MG ions and DDE motif residues, while P1 only made contacts with DNA and IN residues. Conclusion Findings from this study leads to a conclusion that double mutants (S119P, Y143R) and (V150A, M154I) may result in a reduction in the efficacy of DTG, especially when in combination. Furthermore, variants identified in systems P1 and P2 may still allow for effective DTG binding to IN and outcompete viral DNA for host DNA to prevent strand transfer. To the best of our knowledge, this is the first study that uses the consensus WT HIV1C IN sequence to build an accurate 3D homology model to understand the effect of less frequently detected/reported variants on DTG binding in a South African context.
https://etd
Molecular dynamic simulations to investigate the structural impact of known drug resistance mutations on HIV-1C Integrase-Dolutegravir binding
No full textResistance 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. A homology model of HIV-1 subtype C IN was successfully constructed and validated. 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. Also, interaction analysis was performed between neighbouring residues. Three mutations known to be associated with Raltegravir, Elvitegravir and Dolutegravir resistance were selected; E92Q, G140S and Y143R, for molecular dynamics simulations. The structural quality assessment indicated high reliability of the HIV-1C IN tetrameric structure, with more than 90% confidence in modelled regions. Change in free energy for the three mutants indicated different effects, while simulation analysis showed G140S to have the largest affect on protein stability and flexibility.
This was further supported by weaker non-bonded pairwise interaction energy and binding free energy values between the drug DTG and E92Q, Y143R and G140S mutants suggesting reduced binding affinity, as indicated by interaction analysis in comparison to the WT. Our findings suggest the G140S mutant has the strongest effect on the HIV-1C IN protein structure and Dolutegravir binding. To the best of our knowledge, this is the first study that uses the consensus wild type HIV-1C IN sequence to build an accurate 3D model to understand the effect of three known mutations on DTG drug binding in a South Africa context
Interaction analysis of statistically enriched mutations identified in Cameroon recombinant subtype CRF02_AG that can influence the development of Dolutegravir drug resistance mutations
No full textThe 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.We evaluated the HIV-1 CRF02_AG IN gene sequences from Cameroon for the presence of resistanceassociated 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
Abatement of Amoxicillin, Ampicillin, and Chloramphenicol from Aqueous solutions using activated carbon prepared from grape slurry
No full textArticleThe adsorption of amoxicillin (AMX), ampicillin (AMP), and chloramphenicol (CHLR) from simulated antibiotic-contaminated water using adsorbents prepared from grape slurry waste is studied. Batch adsorption experiments are carried out to evaluate the adsorption capacity of the adsorbents for AMX, AMP, and CHLR. Adsorption isotherms are described by the Langmuir and Freundlich isotherms, while the pseudo-second order kinetics describe the sorption processes. Negative values of the enthalpy change show that the sorption processes are exothermic, and the positive values of the Gibbs free energy change indicates non-spontaneous but feasible nature of the adsorption. The study shows that grape slurry waste could be a good precursor to prepare effective adsorbents for the remediation of antibiotic-contaminated wastewater
Molecular dynamic simulations to investigate the structural impact of known drug resistance mutations on HIV-1C Integrase-Dolutegravir binding.
No full textResistance 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. A homology model of HIV-1 subtype C IN was successfully constructed and validated. 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. Also, interaction analysis was performed between neighbouring residues. Three mutations known to be associated with Raltegravir, Elvitegravir and Dolutegravir resistance were selected; E92Q, G140S and Y143R, for molecular dynamics simulations. The structural quality assessment indicated high reliability of the HIV-1C IN tetrameric structure, with more than 90% confidence in modelled regions. Change in free energy for the three mutants indicated different effects, while simulation analysis showed G140S to have the largest affect on protein stability and flexibility. This was further supported by weaker non-bonded pairwise interaction energy and binding free energy values between the drug DTG and E92Q, Y143R and G140S mutants suggesting reduced binding affinity, as indicated by interaction analysis in comparison to the WT. Our findings suggest the G140S mutant has the strongest effect on the HIV-1C IN protein structure and Dolutegravir binding. To the best of our knowledge, this is the first study that uses the consensus wild type HIV-1C IN sequence to build an accurate 3D model to understand the effect of three known mutations on DTG drug binding in a South Africa context
Look East Policy: The Case of Zimbabwe–China Political and Economic Relations Since 2000
No full text This article maps the evolution of Zimbabwe’s Look East Policy (LEP) and specifically the bilateral relationship with China through the lens of Zimbabwe’s domestic politics. It argues that political elite in Zimbabwe has a vested interest in a close economic and political relationship with China at the cost of the interests of the people of Zimbabwe. The author establishes that Zimbabwe’s LEP was intended to respond to the economic sanctions imposed on it by Western nations. From the descriptive account of the LEP provided in the article, it appears that the LEP has been successful in doing that by having a broad-based economic and political relationship with China. The author further critiques the impact of Chinese investment in Zimbabwe as detrimental to the interests of the people. Foreign policy is an instrument that governs and protects the interests of governments, nationals, institutions, organisations and entities within the lenses of bilateral relations between the countries concerned. The Zimbabwe–China relations point to the fact that the latter China is politically and economically committed to engage and develop the former. However, at the heart of commitment and development in Zimbabwe lies questions of interests and the nature of the relationship which is affecting development and commitment to take place. Hence, this article argues that the failure of Zimbabwe to yield satisfying results from the bilateral relations lies mostly on the political and economic weaknesses of the Zimbabwean government and leadership. The fact that the LEP is not formally and publicly developed and disseminated to key stakeholders and the general public and that it remains largely an oral secret public policy statement affects the interests of Zimbabwean economy and interested stakeholders. This also reflects a weak foreign policy directive. As long as Zimbabwe continues to deny to engage with other superpowers and global institutions, the LEP will remain doomed as China will continue to manipulate and exploit the relationship knowingly that Zimbabwe has no other friends and partners for development and cooperation. </jats:p
Interaction analysis of statistically enriched mutations identified in Cameroon recombinant subtype CRF02_AG that can influence the development of Dolutegravir drug resistance mutations
No full textCITATION: Mikasi, S. G., et al. 2021. Interaction analysis of statistically enriched mutations identified in Cameroon recombinant subtype CRF02_AG that can influence the development of Dolutegravir drug resistance mutations. BMC Infectious Diseases, 21:379, doi:10.1186/s12879-021-06059-x.The original publication is available at https://bmcinfectdis.biomedcentral.comBackground: 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 resistanceassociated
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 12.8% (37/287) sequences to contain RAMs, with only 1.0% (3/287) of the sequences having
major INSTI RAMs: T66A, Q148H, R263K and N155H. Of these,11.8% (34/287) of the sequences contained five
different IN accessory mutations; namely Q95K, T97A, G149A, E157Q and D232N. NOPs occurred at a frequency of
66% on the central core domain (CCD) position, 44% on the C-terminal domain (CTD) position and 35% of the Nterminal
domain (NTD) position. The interaction analysis revealed that DTG bound to DNA, 2MG ions and DDE
motif residues for T66A, T97A, Q148H, N155H and R263K comparable to the WT structure. Except for accessory
mutant structure E157Q, only one MG contact was made with DTG, while DTG had no MG ion contacts and no
DDE motif residue contacts for structure D232N.
Conclusions: Our analysis indicated that all RAM’s that resulted in a change in the number of interactions with
encompassing residues does not affect DTG binding, while accessory mutations E157Q and D232N could affect
DTG binding leading to possible DTG resistance. However, further experimental validation is required to validate the
in silico findings of our study.https://bmcinfectdis.biomedcentral.com/articles/10.1186/s12879-021-06059-xPublisher's versio
