Description:
<jats:title>ABSTRACT</jats:title><jats:p>The antigen 85 (Ag85) protein family, consisting of Ag85A, -B, and -C, is vital for<jats:named-content xmlns:xlink="http://www.w3.org/1999/xlink" content-type="genus-species" xlink:type="simple">Mycobacterium tuberculosis</jats:named-content>due to its role in cell envelope biogenesis. The mycoloyl transferase activity of these proteins generates trehalose dimycolate (TDM), an envelope lipid essential for<jats:named-content xmlns:xlink="http://www.w3.org/1999/xlink" content-type="genus-species" xlink:type="simple">M. tuberculosis</jats:named-content>virulence, and cell wall arabinogalactan-linked mycolic acids. Inhibition of these enzymes through substrate analogs hinders growth of mycobacteria, but a link to mycolic acid synthesis has not been established. In this study, we characterized a novel inhibitor of Ag85C, 2-amino-6-propyl-4,5,6,7-tetrahydro-1-benzothiophene-3-carbonitrile (I3-AG85). I3-AG85 was isolated from a panel of four inhibitors that exhibited structure- and dose-dependent inhibition of<jats:named-content xmlns:xlink="http://www.w3.org/1999/xlink" content-type="genus-species" xlink:type="simple">M. tuberculosis</jats:named-content>division in broth culture. I3-AG85 also inhibited<jats:named-content xmlns:xlink="http://www.w3.org/1999/xlink" content-type="genus-species" xlink:type="simple">M. tuberculosis</jats:named-content>survival in infected primary macrophages. Importantly, it displayed an identical MIC against the drug-susceptible H37Rv reference strain and a panel of extensively drug-resistant/multidrug-resistant<jats:named-content xmlns:xlink="http://www.w3.org/1999/xlink" content-type="genus-species" xlink:type="simple">M. tuberculosis</jats:named-content>strains. Nuclear magnetic resonance analysis indicated binding of I3-AG85 to Ag85C, similar to its binding to the artificial substrate octylthioglucoside. Quantification of mycolic acid-linked lipids of the<jats:named-content xmlns:xlink="http://www.w3.org/1999/xlink" content-type="genus-species" xlink:type="simple">M. tuberculosis</jats:named-content>envelope showed a specific blockade of TDM synthesis. This was accompanied by accumulation of trehalose monomycolate, while the overall mycolic acid abundance remained unchanged. Inhibition of Ag85C activity also disrupted the integrity of the<jats:named-content xmlns:xlink="http://www.w3.org/1999/xlink" content-type="genus-species" xlink:type="simple">M. tuberculosis</jats:named-content>envelope. I3-AG85 inhibited the division of and reduced TDM synthesis in an<jats:named-content xmlns:xlink="http://www.w3.org/1999/xlink" content-type="genus-species" xlink:type="simple">M. tuberculosis</jats:named-content>strain deficient in Ag85C. Our results indicate that Ag85 proteins are promising targets for novel antimycobacterial drug design.</jats:p>