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Structural and Functional Characterization of Rv0792c from Mycobacterium tuberculosis: Identifying Small Molecule Inhibitor against HutC Protein
In order to adapt in host tissues, microbial pathogens regulate their gene expression through a variety of transcription factors. Here, we have functionally characterized Rv0792c, a HutC homolog from Mycobacterium tuberculosis. In comparison to the parental strain, a strain of M. tuberculosis with a...
Autores principales: | , , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
American Society for Microbiology
2022
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9927256/ https://www.ncbi.nlm.nih.gov/pubmed/36507689 http://dx.doi.org/10.1128/spectrum.01973-22 |
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author | Chauhan, Neeraj Kumar Anand, Anjali Sharma, Arun Dhiman, Kanika Gosain, Tannu Priya Singh, Prashant Singh, Padam Khan, Eshan Chattopadhyay, Gopinath Kumar, Amit Sharma, Deepak Ashish, Sharma, Tarun Kumar Singh, Ramandeep |
author_facet | Chauhan, Neeraj Kumar Anand, Anjali Sharma, Arun Dhiman, Kanika Gosain, Tannu Priya Singh, Prashant Singh, Padam Khan, Eshan Chattopadhyay, Gopinath Kumar, Amit Sharma, Deepak Ashish, Sharma, Tarun Kumar Singh, Ramandeep |
author_sort | Chauhan, Neeraj Kumar |
collection | PubMed |
description | In order to adapt in host tissues, microbial pathogens regulate their gene expression through a variety of transcription factors. Here, we have functionally characterized Rv0792c, a HutC homolog from Mycobacterium tuberculosis. In comparison to the parental strain, a strain of M. tuberculosis with a Rv0792c mutant was compromised for survival upon exposure to oxidative stress and infection in guinea pigs. RNA sequencing analysis revealed that Rv0792c regulates the expression of genes involved in stress adaptation and virulence of M. tuberculosis. Solution small-angle X-ray scattering (SAXS) data-steered model building confirmed that the C-terminal region plays a pivotal role in dimer formation. Systematic evolution of ligands by exponential enrichment (SELEX) resulted in the identification of single-strand DNA (ssDNA) aptamers that can be used as a tool to identify small-molecule inhibitors targeting Rv0792c. Using SELEX and SAXS data-based modeling, we identified residues essential for Rv0792c’s aptamer binding activity. In this study, we also identified I-OMe-Tyrphostin as an inhibitor of Rv0792c’s aptamer and DNA binding activity. The identified small molecule reduced the growth of intracellular M. tuberculosis in macrophages. The present study thus provides a detailed shape-function characterization of a HutC family of transcription factor from M. tuberculosis. IMPORTANCE Prokaryotes encode a large number of GntR family transcription factors that are involved in various fundamental biological processes, including stress adaptation and pathogenesis. Here, we investigated the structural and functional role of Rv0792c, a HutC homolog from M. tuberculosis. We demonstrated that Rv0792c is essential for M. tuberculosis to adapt to oxidative stress and establish disease in guinea pigs. Using a systematic evolution of ligands by exponential enrichment (SELEX) approach, we identified ssDNA aptamers from a random ssDNA library that bound to Rv0792c protein. These aptamers were thoroughly characterized using biochemical and biophysical assays. Using SAXS, we determined the structural model of Rv0792c in both the presence and absence of the aptamers. Further, using a combination of SELEX and SAXS methodologies, we identified I-OMe-Tyrphostin as a potential inhibitor of Rv0792c. Here we provide a detailed functional characterization of a transcription factor belonging to the HutC family from M. tuberculosis. |
format | Online Article Text |
id | pubmed-9927256 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | American Society for Microbiology |
record_format | MEDLINE/PubMed |
spelling | pubmed-99272562023-02-15 Structural and Functional Characterization of Rv0792c from Mycobacterium tuberculosis: Identifying Small Molecule Inhibitor against HutC Protein Chauhan, Neeraj Kumar Anand, Anjali Sharma, Arun Dhiman, Kanika Gosain, Tannu Priya Singh, Prashant Singh, Padam Khan, Eshan Chattopadhyay, Gopinath Kumar, Amit Sharma, Deepak Ashish, Sharma, Tarun Kumar Singh, Ramandeep Microbiol Spectr Research Article In order to adapt in host tissues, microbial pathogens regulate their gene expression through a variety of transcription factors. Here, we have functionally characterized Rv0792c, a HutC homolog from Mycobacterium tuberculosis. In comparison to the parental strain, a strain of M. tuberculosis with a Rv0792c mutant was compromised for survival upon exposure to oxidative stress and infection in guinea pigs. RNA sequencing analysis revealed that Rv0792c regulates the expression of genes involved in stress adaptation and virulence of M. tuberculosis. Solution small-angle X-ray scattering (SAXS) data-steered model building confirmed that the C-terminal region plays a pivotal role in dimer formation. Systematic evolution of ligands by exponential enrichment (SELEX) resulted in the identification of single-strand DNA (ssDNA) aptamers that can be used as a tool to identify small-molecule inhibitors targeting Rv0792c. Using SELEX and SAXS data-based modeling, we identified residues essential for Rv0792c’s aptamer binding activity. In this study, we also identified I-OMe-Tyrphostin as an inhibitor of Rv0792c’s aptamer and DNA binding activity. The identified small molecule reduced the growth of intracellular M. tuberculosis in macrophages. The present study thus provides a detailed shape-function characterization of a HutC family of transcription factor from M. tuberculosis. IMPORTANCE Prokaryotes encode a large number of GntR family transcription factors that are involved in various fundamental biological processes, including stress adaptation and pathogenesis. Here, we investigated the structural and functional role of Rv0792c, a HutC homolog from M. tuberculosis. We demonstrated that Rv0792c is essential for M. tuberculosis to adapt to oxidative stress and establish disease in guinea pigs. Using a systematic evolution of ligands by exponential enrichment (SELEX) approach, we identified ssDNA aptamers from a random ssDNA library that bound to Rv0792c protein. These aptamers were thoroughly characterized using biochemical and biophysical assays. Using SAXS, we determined the structural model of Rv0792c in both the presence and absence of the aptamers. Further, using a combination of SELEX and SAXS methodologies, we identified I-OMe-Tyrphostin as a potential inhibitor of Rv0792c. Here we provide a detailed functional characterization of a transcription factor belonging to the HutC family from M. tuberculosis. American Society for Microbiology 2022-12-12 /pmc/articles/PMC9927256/ /pubmed/36507689 http://dx.doi.org/10.1128/spectrum.01973-22 Text en Copyright © 2022 Chauhan et al. https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license (https://creativecommons.org/licenses/by/4.0/) . |
spellingShingle | Research Article Chauhan, Neeraj Kumar Anand, Anjali Sharma, Arun Dhiman, Kanika Gosain, Tannu Priya Singh, Prashant Singh, Padam Khan, Eshan Chattopadhyay, Gopinath Kumar, Amit Sharma, Deepak Ashish, Sharma, Tarun Kumar Singh, Ramandeep Structural and Functional Characterization of Rv0792c from Mycobacterium tuberculosis: Identifying Small Molecule Inhibitor against HutC Protein |
title | Structural and Functional Characterization of Rv0792c from Mycobacterium tuberculosis: Identifying Small Molecule Inhibitor against HutC Protein |
title_full | Structural and Functional Characterization of Rv0792c from Mycobacterium tuberculosis: Identifying Small Molecule Inhibitor against HutC Protein |
title_fullStr | Structural and Functional Characterization of Rv0792c from Mycobacterium tuberculosis: Identifying Small Molecule Inhibitor against HutC Protein |
title_full_unstemmed | Structural and Functional Characterization of Rv0792c from Mycobacterium tuberculosis: Identifying Small Molecule Inhibitor against HutC Protein |
title_short | Structural and Functional Characterization of Rv0792c from Mycobacterium tuberculosis: Identifying Small Molecule Inhibitor against HutC Protein |
title_sort | structural and functional characterization of rv0792c from mycobacterium tuberculosis: identifying small molecule inhibitor against hutc protein |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9927256/ https://www.ncbi.nlm.nih.gov/pubmed/36507689 http://dx.doi.org/10.1128/spectrum.01973-22 |
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