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Using a drug-repurposing platform, we identified TI-374, a hydroxamic acid containing compound that inhibits\n                  <jats:italic>Mycobacterium tuberculosis<\/jats:italic>\n                  (\n                  <jats:italic>Mtb<\/jats:italic>\n                  ) with sub-micromolar potency. Systems analysis, resistance mapping, supplementation assays, and biochemical studies showed that TI-374 inhibits two PLP-dependent aminotransferases, AlaA and HisC1. However, its activity is driven primarily by irreversible inhibition of AlaA, whereas HisC1 inhibition is only partially reversible, revealing differential reversibility between the two targets. Optimization yielded TI-801, a low-nanomolar AlaA inhibitor. Both compounds remained active against intracellular\n                  <jats:italic>Mtb<\/jats:italic>\n                  in a macrophage infection model, where alanine supplementation did not rescue growth, indicating that host-derived alanine is unlikely to bypass AlaA inhibition. Genetic deletion of\n                  <jats:italic>alaA<\/jats:italic>\n                  attenuated\n                  <jats:italic>Mtb<\/jats:italic>\n                  survival in a murine infection model. Together, these findings support AlaA as a host-relevant metabolic vulnerability in\n                  <jats:italic>Mtb<\/jats:italic>\n                  and TI-801 as a mechanistic chemical probe for its validation as an antitubercular target.\n                <\/jats:p>","DOI":"10.64898\/2026.06.12.731178","type":"posted-content","created":{"date-parts":[[2026,6,16]],"date-time":"2026-06-16T20:45:20Z","timestamp":1781642720000},"source":"Crossref","is-referenced-by-count":0,"title":["Induced alanine auxotrophy as a therapeutic strategy against\n                  <i>Mycobacterium tuberculosis<\/i>"],"prefix":"10.64898","author":[{"given":"Menna-Allah W.","family":"Shalaby","sequence":"first","affiliation":[{"name":"Borch Department of Medicinal Chemistry and Molecular Pharmacology, College of Pharmacy, Purdue University, West Lafayette, Indiana 47907, United States"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nagabhushana C.","family":"Beeralingappa","sequence":"additional","affiliation":[{"name":"Borch Department of Medicinal Chemistry and Molecular Pharmacology, College of Pharmacy, Purdue University, West Lafayette, Indiana 47907, United States"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Annadka","family":"Shrinidhi","sequence":"additional","affiliation":[{"name":"Borch Department of Medicinal Chemistry and Molecular Pharmacology, College of Pharmacy, Purdue University, West Lafayette, Indiana 47907, United States"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Gaelle Guiewi","family":"Makafe","sequence":"additional","affiliation":[{"name":"Trudeau Institute, Saranac Lake, NY, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Evan","family":"Nece","sequence":"additional","affiliation":[{"name":"Trudeau Institute, Saranac Lake, NY, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Aditya","family":"Patwardhan","sequence":"additional","affiliation":[{"name":"Trudeau Institute, Saranac Lake, NY, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Timothy","family":"Low-Beer","sequence":"additional","affiliation":[{"name":"Trudeau Institute, Saranac Lake, NY, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Atsuo","family":"Kuki","sequence":"additional","affiliation":[{"name":"Trudeau Institute, Saranac Lake, NY, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Felix","family":"Sheinerman","sequence":"additional","affiliation":[{"name":"Trudeau Institute, Saranac Lake, NY, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0880-4487","authenticated-orcid":false,"given":"Brian","family":"Weinrick","sequence":"additional","affiliation":[{"name":"Trudeau Institute, Saranac Lake, NY, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8305-0606","authenticated-orcid":false,"given":"Daniel P.","family":"Flaherty","sequence":"additional","affiliation":[{"name":"Borch Department of Medicinal Chemistry and Molecular Pharmacology, College of Pharmacy, Purdue University, West Lafayette, Indiana 47907, United States"},{"name":"Purdue Institute for Drug Discovery, West Lafayette, IN 47907, United States"},{"name":"Purdue Institute of Inflammation, Immunology and Infectious Disease, West Lafayette, IN 47907, United States"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0245-9918","authenticated-orcid":false,"given":"Michaelle","family":"Chojnacki","sequence":"additional","affiliation":[{"name":"Trudeau Institute, Saranac Lake, NY, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2026061800551115000_2026.06.12.731178v1.1","unstructured":"World Health Organization. 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