Several studies have described FadD, the Escherichia coli fatty acyl-CoA synthetase [also known as fatty acid :CoA ligase (AMPforming) ; EC 6.2.1.3], as a 42-50 kDa enzyme. Based on sequencing and expression data from the fadD gene, other reports have suggested that FadD is a 62 kDa protein and represents the sole fatty acyl-CoA synthetase in E. coli. We report that the 62 kDa FadD enzyme is a substrate for the outer membrane protease OmpT in itro, producing a 43 kDa Cterminal fragment and a 19 kDa N-terminal fragment. Immunoblotting with a FadD antibody revealed that only the 62 kDa form of the enzyme is present in i o, but we utilized the proteolytic sensitivity of FadD to investigate its structure. Photoaffinity labelling experiments revealed that both intact FadD and the 43 kDa fragment bound a long-chain fatty acid.
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