Zainab Edoo, Astrid Lenne-Delmotte, Camille Grosse, Marie Devaere, Marion Michel, Guillaume Caron, Ernesto Anoz-Carbonell, Kamel Djaout, Rosangela Frita, Cyril Gaudin, Line Hofmann, Sophie Lecher, Véronique Megalizzi, Alessia Michelotti, David Rengel, Pauline Rouan, Stephanie Slupek, Lina Tawk, Rudy Antoine, Hanna Kulyk, Glenn Dale, Christophe Guilhot, Nicolas Willand, Guy Lippens, René Wintjens, Alain R Baulard
Tuberculosis remains the leading cause of death from a single infectious agent, and rising multi-drug resistance in Mycobacterium tuberculosis (Mtb) underscores the urgent need for new antibiotics. Here, we characterize BVL3572S, a hydroxamic acid-containing compound that is bactericidal against Mtb. The primary targets of BVL3572S are the pyridoxal phosphate (PLP; active form of vitamin B6)-dependent aminotransferases HisC (Rv1600) and AlaA (Rv0337c; formerly AspC), simultaneously impacting L-histidine and L-alanine biosynthesis. X-ray crystallography revealed a covalent PLP-BVL3572S adduct within the HisC active site. The formation of the adduct followed by its release suggests a futile cycle that depletes PLP. Consistently, isotopic labeling revealed widespread perturbation of amino acid biosynthesis. CRISPRi and Tn-seq analyses additionally indicated disruptions in central metabolism, cell envelope integrity, and redox balance. BVL3572S displayed strong synergy with the antitubercular drug D-cycloserine. Collectively, our findings establish BVL3572S as a promising lead compound acting through a previously unexploited, multitarget mechanism.