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The emergence of\n                    <jats:italic>Mtb<\/jats:italic>\n                    's multidrug\u2010resistant and extremely drug\u2010resistant strains has imposed a great challenge for TB treatment. Hence, there is always a demand to explore new targets that may be crucial for the survival and pathogenicity of the bacilli. Oxidoreductases are a class of enzymes that transfer electrons in various biological pathways and reactions, at the expense of cellular NADPH\/NADH. Here, we analyzed oxidoreductases from the H37Rv proteome and identified two uncharacterized putative oxidoreductases, Rv1260 and Rv1714. These putative oxidoreductases showed conservation among pathogenic and opportunistic mycobacterial species and were predicted to be virulence factors essential for the pathogen's survival. The 3D structural model and amino acid sequence of one of the oxidoreductases, Rv1260, showed similarities with tetracycline destructase, a flavin\u2010dependent monooxygenase. Thin\u2010layer chromatography and UV\u2013visible spectroscopic experiments confirmed the presence of the FAD molecule in a bound form with the recombinant protein. Fluorescence quenching studies demonstrated a comparatively better affinity of NADPH than NADH with the protein. The protein also displayed efficient binding with chlortetracycline. Molecular dynamics simulations were employed to gain insights into the substrate binding and conformational changes in the protein. Moreover, the importance of the substrate binding region, the C\u2010terminal helix, and the FAD binding cavity, located near the isoalloxazine ring, was highlighted. Overall, the study provides biochemical, biophysical, and mechanistic insights into one of the putative\n                    <jats:italic>Mtb<\/jats:italic>\n                    oxidoreductases. Based on our data, we propose that this protein may perform monooxygenation functions under specific redox conditions and contribute to the redox processes in\n                    <jats:italic>Mtb<\/jats:italic>\n                    .\n                  <\/jats:p>","DOI":"10.1002\/prot.70095","type":"journal-article","created":{"date-parts":[[2025,12,6]],"date-time":"2025-12-06T04:44:28Z","timestamp":1764996268000},"page":"999-1016","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Biochemical, Biophysical, and Mechanistic Insights on a Putative Oxidoreductase From\n                    <scp>\n                      <i>Mycobacterium tuberculosis<\/i>\n                    <\/scp>\n                    That Shares Structural Homology With Tetracycline Destructase"],"prefix":"10.1002","volume":"94","author":[{"given":"Mohd.","family":"Shiraz","sequence":"first","affiliation":[{"name":"Laboratory of Structural Biology, Department of Biochemistry, School of Life Sciences University of Hyderabad  Gachibowli Hyderabad India"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dipanwita","family":"Biswas","sequence":"additional","affiliation":[{"name":"Laboratory of Structural Biology, Department of Biochemistry, School of Life Sciences University of Hyderabad  Gachibowli Hyderabad India"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"G.","family":"Priyanka","sequence":"additional","affiliation":[{"name":"Department of Biotechnology and Bioinformatics, School of Life Sciences University of Hyderabad  Gachibowli Hyderabad India"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3024-1142","authenticated-orcid":false,"given":"N. Prakash","family":"Prabhu","sequence":"additional","affiliation":[{"name":"Department of Biotechnology and Bioinformatics, School of Life Sciences University of Hyderabad  Gachibowli Hyderabad India"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8462-7166","authenticated-orcid":false,"given":"Mohd.","family":"Akif","sequence":"additional","affiliation":[{"name":"Laboratory of Structural Biology, Department of Biochemistry, School of Life Sciences University of Hyderabad  Gachibowli Hyderabad India"},{"name":"Faculty of Pharmaceutical Sciences UCSI University  Kuala Lumpur 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