A short-chain acyl-CoA synthetase that supports branched-chain fatty acid synthesis in Staphylococcus aureus

• Published in: The Journal of biological chemistry Vol. 299; no. 4; p. 103036
• Main Authors: Whaley, Sarah G., Frank, Matthew W., Rock, Charles O.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.04.2023; American Society for Biochemistry and Molecular Biology
• Subjects: acyl-CoA synthetase; branched-chain ketoacid dehydrogenase; fatty acid synthesis; Fatty Acids - metabolism; Isobutyrates; Ligases; membrane; phospholipid; Staphylococcus aureus; Staphylococcus aureus - genetics; Staphylococcus aureus - metabolism; FASII; MbcS; BHI; membrane; BCFA; HSS; MRM; branched-chain ketoacid dehydrogenase; ACP; UFLC; TBA; fatty acid synthesis; UPLC; PG; acyl-CoA synthetase; Buk; Bkd; phospholipid; Staphylococcus aureus; BCAA; Ptb
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/j.jbc.2023.103036

Staphylococcus aureus controls its membrane biophysical properties using branched-chain fatty acids (BCFAs). The branched-chain acyl-CoA precursors, utilized to initiate fatty acid synthesis, are derived from branched-chain ketoacid dehydrogenase (Bkd), a multiprotein complex that converts α-keto acids to their corresponding acyl-CoAs; however, Bkd KO strains still contain BCFAs. Here, we show that commonly used rich medias contain substantial concentrations of short-chain acids, like 2-methylbutyric and isobutyric acids, that are incorporated into membrane BCFAs. Bkd-deficient strains cannot grow in defined medium unless it is supplemented with either 2-methylbutyric or isobutyric acid. We performed a screen of candidate KO strains and identified the methylbutyryl-CoA synthetase (mbcS gene; SAUSA300_2542) as required for the incorporation of 2-methylbutyric and isobutyric acids into phosphatidylglycerol. Our mass tracing experiments show that isobutyric acid is converted to isobutyryl-CoA that flows into the even-chain acyl-acyl carrier protein intermediates in the type II fatty acid biosynthesis elongation cycle. Furthermore, purified MbcS is an ATP-dependent acyl-CoA synthetase that selectively catalyzes the activation of 2-methylbutyrate and isobutyrate. We found that butyrate and isovalerate are poor MbcS substrates and activity was not detected with acetate or short-chain dicarboxylic acids. Thus, MbcS functions to convert extracellular 2-methylbutyric and isobutyric acids to their respective acyl-CoAs that are used by 3-ketoacyl-ACP synthase III (FabH) to initiate BCFA biosynthesis.