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

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 de-hydrogenase (Bkd), a multiprotein complex that converts alpha-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 methyl-butyryl-CoA synthetase (mbcS gene; SAUSA300_2542) as required for the incorporation of 2-methylbutyric and iso-butyric acids into phosphatidylglycerol. Our mass tracing ex-periments 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.

Automated summary

Staphylococcus aureus uses branched-chain fatty acids (BCFAs) to control the properties of its membrane. Short-chain acids, such as 2-methylbutyric and isobutyric acids, are found in commonly used rich medias and incorporated into membrane BCFAs. The methyl-butyryl-CoA synthetase (MbcS) gene is required for converting these acids into their corresponding acyl-CoAs.