ACCase 6 is the essential acetyl-CoA carboxylase involved in fatty acid and mycolic acid biosynthesis in mycobacteria

Abstract

Mycolic acids are essential for the survival, virulence and antibiotic resistance of the human pathogen <em>Mycobacterium tuberculosis</em>. Inhibitors of mycolic acid biosynthesis, such as isoniazid and ethionamide, have been used as efficient drugs for the treatment of tuberculosis. However, the increase in cases of multidrug-resistant tuberculosis has prompted a search for new targets and agents that could also affect synthesis of mycolic acids. In mycobacteria, the acyl-CoA carboxylases (ACCases) provide the building blocks for <em>de novo</em> fatty acid biosynthesis by fatty acid synthase (FAS) I and for the elongation of FAS I products by the FAS II complex to produce meromycolic acids. By generating a conditional mutant in the <em>accD6</em> gene of <em>Mycobacterium smegmatis,</em> we demonstrated that AccD6 is the essential carboxyltransferase component of the ACCase 6 enzyme complex implicated in the biosynthesis of malonyl-CoA, the substrate of the two FAS enzymes of <em>Mycobacterium</em> species. Based on the conserved structure of the AccD5 and AccD6 active sites we screened several inhibitors of AccD5 as potential inhibitors of AccD6 and found that the ligand NCI-172033 was capable of inhibiting AccD6 with an IC₅₀ of 8 ìM. The compound showed bactericidal activity against several pathogenic <em>Mycobacterium</em> species by producing a strong inhibition of both fatty acid and mycolic acid biosynthesis at minimal inhibitory concentrations. Overexpression of <em>accD6</em> in <em>M. smegmatis</em> conferred resistance to NCI-172033, confirming AccD6 as the main target of the inhibitor. These results define the biological role of a key ACCase in the biosynthesis of membrane and cell envelope fatty acids, and provide a new target, AccD6, for rational development of novel anti-mycobacterial drugs