Microbial fatty acid biosynthetic enzymes are important targets for antibiotic development, chemical manufacturing and biofuel production. In Escherichia coli, the acyl carrier protein (AcpP) plays a central role by sequestering and shuttling the growing acyl chain between fatty acid biosynthetic enzymes. FabA, a β-hydroxylacyl-AcpP dehydratase, is an important enzyme in controlling fatty acid chain length and unsaturated fatty acid amounts. Understanding the 10-carbon chain length preference of FabA, and the preference of other fatty acid biosynthetic enzymes, is crucial for understanding and ultimately manipulating this metabolic pathway. In this study, four mechanistic crosslinking probes mimicking varying acyl chain lengths were synthesized to probe for chain length specificity of fourteen FabA mutants. The three FabA regions mutated were the AcpP interacting “positive patch,” substrate-binding pocket and “gating residues.” These studies shed light on the importance of the “positive patch” for AcpP-partner protein interactions, and help elucidate the chain length preference of FabA.

Using mechanistic crosslinkers to study the substrate selectivity and protein-protein interactions of the E. coli fatty acid dehydratase, FabA

251st National Meeting (2016)

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