Novel Antimicrobials from Uncultured Bacteria Acting against Mycobacterium tuberculosis

, which causes tuberculosis (TB), is estimated to infect one-third of the world's population. The overall burden and the emergence of drug-resistant strains of underscore the need for new therapeutic options against this important human pathogen. Our recent work demonstrated the success of natural product discovery in identifying novel compounds with efficacy against Here, we improve on these methods by combining improved isolation and selective screening to identify three new anti-TB compounds: streptomycobactin, kitamycobactin, and amycobactin. We were unable to obtain mutants resistant to streptomycobactin, and its target remains to be elucidated. We identify the target of kitamycobactin to be the mycobacterial ClpP1P2C1 protease and confirm that kitamycobactin is an analog of the previously identified compound lassomycin. Further, we identify the target of amycobactin to be the essential protein secretion pore SecY. We show further that amycobactin inhibits protein secretion via the SecY translocon. Importantly, this inhibition is bactericidal to nonreplicating This is the first compound, to our knowledge, that targets the Sec protein secretion machinery in This work underscores the ability of natural product discovery to deliver not only new compounds with activity against but also compounds with novel targets. Decreasing discovery rates and increasing resistance have underscored the need for novel therapeutic options to treat infection. Here, we screen extracts from previously uncultured soil microbes for specific activity against , identifying three novel compounds. We further define the mechanism of action of one compound, amycobactin, and demonstrate that it inhibits protein secretion through the Sec translocation machinery.