The unusual convergence of steroid catabolic pathways in Mycobacterium abscessus

Abstract

Mycobacterium abscessus , an opportunistic pathogen responsible for pulmonary infections, contains genes predicted to encode two steroid catabolic pathways: a cholesterol catabolic pathway similar to that of Mycobacterium tuberculosis and a 4-androstenedione (4-AD) catabolic pathway. Consistent with this prediction, M. abscessus grew on both steroids. In contrast to M. tuberculosis , Rhodococcus jostii RHA1, and other Actinobacteria, the cholesterol and 4-AD catabolic gene clusters of the M. abscessus complex lack genes encoding HsaD, the meta -cleavage product (MCP) hydrolase. However, M. abscessus ATCC 19977 harbors two hsaD homologs elsewhere in its genome. Only one of the encoded enzymes detectably transformed steroid metabolites. Among tested substrates, HsaD Mab and HsaD Mtb of M. tuberculosis had highest substrate specificities for MCPs with partially degraded side chains thioesterified with coenzyme A ( k cat / K M = 1.9 × 10 4 and 5.7 × 10 3 mM −1 s −1 , respectively). Consistent with a dual role in cholesterol and 4-AD catabolism, HsaD Mab also transformed nonthioesterified substrates efficiently, and a Δ hsaD mutant of M. abscessus grew on neither steroid. Interestingly, both steroids prevented growth of the mutant on acetate. The Δ hsaD mutant of M. abscessus excreted cholesterol metabolites with a fully degraded side chain, while the corresponding RHA1 mutant excreted metabolites with partially degraded side chains. Finally, the Δ hsaD mutant was not viable in macrophages. Overall, our data establish that the cholesterol and 4-AD catabolic pathways of M. abscessus 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.

Cholesterol-Mediated Coenzyme A Depletion in Catabolic Mutants of Mycobacteria Leads to Toxicity

“…As noted above, disruption of HsaD, a ring A/B-degrading enzyme, appears to be toxic in M. abscessus due to the accumulation of a meta -cleavage product and the sequestration of CoASH . Ultimately, compounds that target cholesterol catabolism may be much more potent in combination with other TB therapeutics.…”

“…Notably, an hsaD mutant in M. abscessus accumulates a steroid-derived metabolite that has a CoA-independent bacteriotoxic effect . However, the toxicity of coumaroyl-CoAs in RHA1, E.…”

“…In the case of the Δ ipdAB mutant, the strain was inoculated in cholesterol media to a final OD 600 of 0.4. Intracellular metabolites were extracted from pelleted cells using 700 μL of solvent as described previously …”

“…In the lower pathway, regulated by KstR2, another TetR-type transcriptional repressor, , HIP catabolism is initiated by FadD3, an acyl-CoA synthetase which thioesterifies HIP to produce HIP-CoA, the inducer for the KstR2 regulon. , The catabolism of HIP-CoA proceeds through a series of acyl-CoA metabolites, and is predicted to yield propionyl-CoA, acetyl-CoA, and succinyl-CoA (Figure ). Organisms in which this pathway has been validated include Mtb, Mycobacterium smegmatis, Mycobacterium abscessus, and Rhodococcus jostii RHA1 (RHA1), a related bacterium that also produces mycolic acids. ,− …”

Unravelling and engineering an operon involved in the side-chain degradation of sterols in Mycolicibacterium neoaurum for the production of steroid synthons

“…Therefore, the degradation process of the C17 side chain of sterols is essential for the production of C-19 or C-22 metabolites [ 29 , 30 ]. The degradation of the C17 side chain is similar to the β-oxidative degradation of fatty acids, and the conversion process from sterols to C-19 steroids is known as the AD sub-pathway [ 31 , 32 ]. The key genes involved in the AD sub-pathway are mainly located in StCat-GC.…”