Identification of selective novel hits against Mycobacterium tuberculosis KasA potential allosteric sites using bioinformatics approaches
- Authors: Hare, Fadzayi Faith
- Date: 2022-10-14
- Subjects: Tuberculosis , Docking , Molecules Models , Virtual screening , Multidrug-resistant tuberculosis , Fatty acids Synthesis , Drugs Design
- Language: English
- Type: Academic theses , Master's theses , text
- Identifier: http://hdl.handle.net/10962/362842 , vital:65367
- Description: Tuberculosis (TB) is a global health threat that has led to approximately 1.5 million deaths annually. According to the World Health Organization (WHO), TB is among the top ten deadly diseases and is the leading cause of death due to a single infectious agent. The main challenge in the effective treatment and control of TB is the ongoing emergence of resistant strains of Mycobacterium tuberculosis (Mtb) which lead to multi-drug resistant (MDR) and extensive-drug resistant (XDR) TB. Hence, the identification and characterization of novel drug targets and drugs that modulate the activity of the pathogen are an urgent priority. The current situation even necessitates the reengineering or repurposing of drugs in order to achieve effective control. The β-ketoacyl-acyl carrier protein synthase I (KasA) of Mycobacterium tuberculosis is an essential enzyme in the mycobacterial fatty acid synthesis (FAS-II) pathway and is believed to be a promising target for drug discovery in TB. It is one of the five main proteins of the FAS-II pathway and catalyzes a key condensation reaction in the synthesis of meromycolate chains, the precursors of mycolic acids involved in cell wall formation. Although this protein has been extensively studied, little research has been devoted to the allosteric inhibition of potential drug compounds. The main aim of this research was to identify the allosteric sites on the protein that could be involved in the inhibition of substrate binding activities and novel drug compounds that bind to these sites by use of in-silico approaches. The bioinformatics approaches used in this study were divided into four main objectives namely identification of KasA homolog sequences, sequence analysis and protein characterization, allosteric site search and lastly virtual screening of DrugBank compounds via molecular docking. Fifteen homolog sequences were identified from the BLASTP analysis and were derived from bacteria, fungi and mammals. In order to discover important residues and regions within the KasA proteins, sequence alignment, motif analysis and phylogenetic studies were performed using Mtb KasA as a reference. Sequence alignment revealed conserved residues in all KasA proteins that have functional importance such as the catalytic triad residues (Cys171, His311 and His345). Motif analysis identified 18 highly conserved motifs within the KasA proteins with structural and functional roles. In addition, motifs unique to the Mtb KasA protein were also identified and explored for inhibitor drug design purposes. Phylogenetic analysis of the homolog sequences showed a distinct clustering of prokaryotes and eukaryotes. A distinctive clustering was also observed for species belonging to the same genus. Since the mechanism of action of most drugs involves the active site, allosteric site search was conducted on Mtb KasA and the human homolog protein using a combination of pocket detection algorithms with the aim of identifying sites that could be utilized in allosteric modulator drug discovery. This was followed by the virtual screening of 2089 FDA approved DrugBank compounds against the entire protein surfaces of Mtb KasA and Hsmt KasA, performed via molecular docking using AutoDock Vina. Screening of the compounds was based on the binding energies, with more focus on identifying ligands that bound exclusively to the acyl-binding tunnel of Mtb KasA. This reduced the data set to 27 promising drug compounds with a relatively high binding affinity for Mtb KasA, however, further experiments need to be performed to validate this result. Among these compounds were DB08889, DB06755, DB09270, DB11226, DB00392, DB12278, DB08936, DB00781, DB13720 and DB00392, which displayed relatively low binding energies for Mtb KasA when compared to the human homolog protein. , Thesis (MSc) -- Faculty of Science, Biochemistry and Microbiology, 2022
- Full Text:
- Date Issued: 2022-10-14
- Authors: Hare, Fadzayi Faith
- Date: 2022-10-14
- Subjects: Tuberculosis , Docking , Molecules Models , Virtual screening , Multidrug-resistant tuberculosis , Fatty acids Synthesis , Drugs Design
- Language: English
- Type: Academic theses , Master's theses , text
- Identifier: http://hdl.handle.net/10962/362842 , vital:65367
- Description: Tuberculosis (TB) is a global health threat that has led to approximately 1.5 million deaths annually. According to the World Health Organization (WHO), TB is among the top ten deadly diseases and is the leading cause of death due to a single infectious agent. The main challenge in the effective treatment and control of TB is the ongoing emergence of resistant strains of Mycobacterium tuberculosis (Mtb) which lead to multi-drug resistant (MDR) and extensive-drug resistant (XDR) TB. Hence, the identification and characterization of novel drug targets and drugs that modulate the activity of the pathogen are an urgent priority. The current situation even necessitates the reengineering or repurposing of drugs in order to achieve effective control. The β-ketoacyl-acyl carrier protein synthase I (KasA) of Mycobacterium tuberculosis is an essential enzyme in the mycobacterial fatty acid synthesis (FAS-II) pathway and is believed to be a promising target for drug discovery in TB. It is one of the five main proteins of the FAS-II pathway and catalyzes a key condensation reaction in the synthesis of meromycolate chains, the precursors of mycolic acids involved in cell wall formation. Although this protein has been extensively studied, little research has been devoted to the allosteric inhibition of potential drug compounds. The main aim of this research was to identify the allosteric sites on the protein that could be involved in the inhibition of substrate binding activities and novel drug compounds that bind to these sites by use of in-silico approaches. The bioinformatics approaches used in this study were divided into four main objectives namely identification of KasA homolog sequences, sequence analysis and protein characterization, allosteric site search and lastly virtual screening of DrugBank compounds via molecular docking. Fifteen homolog sequences were identified from the BLASTP analysis and were derived from bacteria, fungi and mammals. In order to discover important residues and regions within the KasA proteins, sequence alignment, motif analysis and phylogenetic studies were performed using Mtb KasA as a reference. Sequence alignment revealed conserved residues in all KasA proteins that have functional importance such as the catalytic triad residues (Cys171, His311 and His345). Motif analysis identified 18 highly conserved motifs within the KasA proteins with structural and functional roles. In addition, motifs unique to the Mtb KasA protein were also identified and explored for inhibitor drug design purposes. Phylogenetic analysis of the homolog sequences showed a distinct clustering of prokaryotes and eukaryotes. A distinctive clustering was also observed for species belonging to the same genus. Since the mechanism of action of most drugs involves the active site, allosteric site search was conducted on Mtb KasA and the human homolog protein using a combination of pocket detection algorithms with the aim of identifying sites that could be utilized in allosteric modulator drug discovery. This was followed by the virtual screening of 2089 FDA approved DrugBank compounds against the entire protein surfaces of Mtb KasA and Hsmt KasA, performed via molecular docking using AutoDock Vina. Screening of the compounds was based on the binding energies, with more focus on identifying ligands that bound exclusively to the acyl-binding tunnel of Mtb KasA. This reduced the data set to 27 promising drug compounds with a relatively high binding affinity for Mtb KasA, however, further experiments need to be performed to validate this result. Among these compounds were DB08889, DB06755, DB09270, DB11226, DB00392, DB12278, DB08936, DB00781, DB13720 and DB00392, which displayed relatively low binding energies for Mtb KasA when compared to the human homolog protein. , Thesis (MSc) -- Faculty of Science, Biochemistry and Microbiology, 2022
- Full Text:
- Date Issued: 2022-10-14
Exploring socio-economic factors influencing incidences and outcome of multidrug resistance tuberculosis among patients and facility staffs in Makana Sub-District, Eastern Cape
- Cannon, Lesley-Ann https://orcid.org/0000-0002-7635-277X
- Authors: Cannon, Lesley-Ann https://orcid.org/0000-0002-7635-277X
- Date: 2022-02
- Subjects: Multidrug resistance , Tuberculosis
- Language: English
- Type: Master's theses , text
- Identifier: http://hdl.handle.net/10353/26706 , vital:65958
- Description: Background Drug-resistant Tuberculosis (DR-TB) is one of the main causes of global public health crisis, due to the morbidity and mortality rates associated with the disease. This DR TB is a complex illness having direct and indirect impact on finances, social functioning, and quality of life of infected individuals. Major research advances have been made in the diagnosis and treatment of DR-TB. However, minimal information exists on the socio-economic factors influencing the incidence and outcomes. This study aims to fill the gap by exploring the socio-economic factors from both the health care professional and patient perspective in particular settings to gain insights into developing context-specific strategies against the burden of DR-TB. Methodology The study applied a qualitative method to explore the socio-economic factors influencing MDR-TB through key-in-depth interviews (KIIs) and focus group discussions (FGDs). The study enrolled a total of thirty-two (32) consenting participants. The KIIs was conducted for ten (10) healthcare workers and nine (9) MDR-TB patients. Two focus group discussions were done involving seven (7) MDR TB patients and six (6) MDR-TB patients, respectively. The study targeted healthcare workers working in the MDR-TB field and TB patients with the following: GeneXpert Rifampicin resistance and patient confirmed as MDR TB. Eligible participants were selected using a purposive sampling technique from the hospitals` routine data electronic records (EDR-WEB database) and hardcopy registers (drug-resistant TB register) on MDR-TB patients enrolled in care at the study site. Informed consent was obtained from all study participants after thoroughly explaining the purpose. No personal information of participants was used. All responses from respondents were coded during analysis for autonomy and the respondents were not identifiable in any published or unpublished work following this research. The interviews were transcribed, some translated into English, where necessary, and analysed until saturation was reached. Data was coded and analysed using both thematic and content analysis technique. Results There were 3 main themes identified in the study: social factors, economic factors, and other contributing factors. 7 sub- themes were recorded under social factors and 2 subthemes under economic factors. Two independent factors that were also considered to impact MDR-TB were the attitude of healthcare workers, as well as the current COVID-19 pandemic. Conclusion MDR-TB is a major public health concern in the Makana Sub-district of the Eastern Cape. The findings of this study highlight the impact of socio- economic factors on the incidence, spread, defaulter rate and outcomes of MDR-TB. The social areas highlighted by the study participants as affecting the incidence and outcomes of MDR TB were housing and relocation, decreased immunity, stigma, patients’ attitude and lack of support, alcohol and other substance usage and prison/ incarceration. The economic factors identified by the participants were unemployment and job loss and health related expenses. Other factors are those factors contributing to the increased incidence and possible poor outcomes of MDR TB. Healthcare workers impact and attitude and the effects of the covid-19 pandemic were highlighted as additional factors influencing the incidence and outcomes of MDR TB. The management of MDR-TB requires rigorous efforts that should be directed at addressing the socio-economic factors. Therefore, future quantitative studies and important programmatic strategies should be considered to tackle the socio-economic challenges that contribute to the burden of MDR-TB infection in the Makana community. , Thesis (MPA) -- Faculty of Health Sciences, 2022
- Full Text:
- Date Issued: 2022-02
- Authors: Cannon, Lesley-Ann https://orcid.org/0000-0002-7635-277X
- Date: 2022-02
- Subjects: Multidrug resistance , Tuberculosis
- Language: English
- Type: Master's theses , text
- Identifier: http://hdl.handle.net/10353/26706 , vital:65958
- Description: Background Drug-resistant Tuberculosis (DR-TB) is one of the main causes of global public health crisis, due to the morbidity and mortality rates associated with the disease. This DR TB is a complex illness having direct and indirect impact on finances, social functioning, and quality of life of infected individuals. Major research advances have been made in the diagnosis and treatment of DR-TB. However, minimal information exists on the socio-economic factors influencing the incidence and outcomes. This study aims to fill the gap by exploring the socio-economic factors from both the health care professional and patient perspective in particular settings to gain insights into developing context-specific strategies against the burden of DR-TB. Methodology The study applied a qualitative method to explore the socio-economic factors influencing MDR-TB through key-in-depth interviews (KIIs) and focus group discussions (FGDs). The study enrolled a total of thirty-two (32) consenting participants. The KIIs was conducted for ten (10) healthcare workers and nine (9) MDR-TB patients. Two focus group discussions were done involving seven (7) MDR TB patients and six (6) MDR-TB patients, respectively. The study targeted healthcare workers working in the MDR-TB field and TB patients with the following: GeneXpert Rifampicin resistance and patient confirmed as MDR TB. Eligible participants were selected using a purposive sampling technique from the hospitals` routine data electronic records (EDR-WEB database) and hardcopy registers (drug-resistant TB register) on MDR-TB patients enrolled in care at the study site. Informed consent was obtained from all study participants after thoroughly explaining the purpose. No personal information of participants was used. All responses from respondents were coded during analysis for autonomy and the respondents were not identifiable in any published or unpublished work following this research. The interviews were transcribed, some translated into English, where necessary, and analysed until saturation was reached. Data was coded and analysed using both thematic and content analysis technique. Results There were 3 main themes identified in the study: social factors, economic factors, and other contributing factors. 7 sub- themes were recorded under social factors and 2 subthemes under economic factors. Two independent factors that were also considered to impact MDR-TB were the attitude of healthcare workers, as well as the current COVID-19 pandemic. Conclusion MDR-TB is a major public health concern in the Makana Sub-district of the Eastern Cape. The findings of this study highlight the impact of socio- economic factors on the incidence, spread, defaulter rate and outcomes of MDR-TB. The social areas highlighted by the study participants as affecting the incidence and outcomes of MDR TB were housing and relocation, decreased immunity, stigma, patients’ attitude and lack of support, alcohol and other substance usage and prison/ incarceration. The economic factors identified by the participants were unemployment and job loss and health related expenses. Other factors are those factors contributing to the increased incidence and possible poor outcomes of MDR TB. Healthcare workers impact and attitude and the effects of the covid-19 pandemic were highlighted as additional factors influencing the incidence and outcomes of MDR TB. The management of MDR-TB requires rigorous efforts that should be directed at addressing the socio-economic factors. Therefore, future quantitative studies and important programmatic strategies should be considered to tackle the socio-economic challenges that contribute to the burden of MDR-TB infection in the Makana community. , Thesis (MPA) -- Faculty of Health Sciences, 2022
- Full Text:
- Date Issued: 2022-02
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