The unusual convergence of steroid catabolic pathways in <i>Mycobacterium abscessus</i>

Abstract

<i>Mycobacterium abscessus</i>, an opportunistic pathogen responsible for pulmonary infections, contains genes predicted to encode two steroid catabolic pathways: a cholesterol catabolic pathway similar to that of <i>Mycobacterium tuberculosis</i> and a 4-androstenedione (4-AD) catabolic pathway. Consistent with this prediction, <i>M. abscessus</i> grew on both steroids. In contrast to <i>M. tuberculosis</i>, <i>Rhodococcus jostii</i> RHA1, and other Actinobacteria, the cholesterol and 4-AD catabolic gene clusters of the <i>M. abscessus</i> complex lack genes encoding HsaD, the <i>meta</i>-cleavage product (MCP) hydrolase. However, <i>M. abscessus</i> ATCC 19977 harbors two <i>hsaD</i> homologs elsewhere in its genome. Only one of the encoded enzymes detectably transformed steroid metabolites. Among tested substrates, HsaD_Mab and HsaD_Mtb of <i>M. tuberculosis</i> had highest substrate specificities for MCPs with partially degraded side chains thioesterified with coenzyme A (<i>k</i>_cat/<i>K</i>_M = 1.9 × 10⁴ and 5.7 × 10³ mM^-1s^-1, respectively). Consistent with a dual role in cholesterol and 4-AD catabolism, HsaD_Mab also transformed nonthioesterified substrates efficiently, and a Δ<i>hsaD</i> mutant of <i>M. abscessus</i> grew on neither steroid. Interestingly, both steroids prevented growth of the mutant on acetate. The Δ<i>hsaD</i> mutant of <i>M. abscessus</i> excreted cholesterol metabolites with a fully degraded side chain, while the corresponding RHA1 mutant excreted metabolites with partially degraded side chains. Finally, the Δ<i>hsaD</i> mutant was not viable in macrophages. Overall, our data establish that the cholesterol and 4-AD catabolic pathways of <i>M. abscessus</i> are unique in that they converge upstream of where this occurs in characterized steroid-catabolizing bacteria. The data further indicate that cholesterol is a substrate for intracellular bacteria and that cholesterol-dependent toxicity is not strictly dependent on coenzyme A sequestration.