Stereochemical Course of Enoyl Reduction Catalyzed by Fatty Acid Synthetase1Stereochemistry of Hydrogen Incorporation from Reduced Pyridine Nucleotide

Abstract

The stereochemical course of the enoyl reduction catalyzed by fatty acid synthetase was investigated using the enzymes from rat liver and Brevibacterium ammoniagenes. Deuterium-labeled fatty acids were synthesized by incubating the synthetases with either 4R[4-²H₂]- or 4S-[4-²H₁]NADPH. The deuterium-labeled fatty acids thus produced were subjected to the action of a stereospecific enzyme, acyl-CoA oxidase. The deuterium-labeled fatty acids and 2,3-dehydroacyl thioesters, the products of acyl-CoA oxidase, were methylated and analyzed for deuterium content by gas chromatography-mass spectrometry. These experiments provided information to determine the configuration at the C-3 position of fatty acids formed by fatty acid synthetases. The results suggested that the stereochemistry of hydrogen (as hydride) incorporation from reduced pyridine nucleotides during enoyl reduction was different between rat liver and B. ammoniagenes synthetases: the enoyl reduction of rat liver enzyme involved the re-attack of hydride and that of B. ammoniagenes enzyme involved the si-attack of hydride.²