IN-VIVO AND IN-VITRO EFFECTS OF THIOLACTOMYCIN ON FATTY-ACID BIOSYNTHESIS IN STREPTOMYCES-COLLINUS

A stable-isotope assay was used to analyze the effectiveness of variou s perdeuterated short-chain acyl coenzyme A (acyl-CoA) compounds as st arter units for straight- and branched-chain fatty acid biosynthesis i n cell extracts of Streptomyces collinus. In these extracts perdeutera ted isobutyryl-CoA was converted to isopalmitate (a branched-chain fat ty acid), while butyryl-CoA was converted to palmitate (a straight-cha in fatty acid), These observations are consistent with precious in viv o analyses of fatty acid biosynthesis in S. collinus, which suggested that butyryl-CoA and isobutyryl-CoA function as starter units for palm itate and isopalmitate biosynthesis, respectively, Additionally, in vi tro analysis demonstrated that acetyl-CoA can function as a starter un it for palmitate biosynthesis. Palmitate biosynthesis and isopalmitate biosynthesis in these cell extracts were both effectively inhibited b y thiolactomycin, a known type II fatty acid synthase inhibitor, In vi vo experiments demonstrated that concentrations of thiolactomycin rang ing from 0.1 to 0.2 mg/ml produced both a dramatic decrease in the cel lular levels of branched-chain fatty acids and a surprising three- to fivefold increase in the cellular levels of the straight-chain fatty a cids palmitate and myristate. Additional in vivo incorporation studies with perdeuterated butyrate suggested that, in accord with the in vit ro studies, the biosynthesis of the palmitate from butyryl-CoA decreas es in the presence of thiolactomycin. In contrast, in vivo incorporati on studies with perdeuterated acetate demonstrated that the biosynthes is of palmitate from acetyl-CoA increases in the presence of thiolacto mycin. These observations clearly demonstrate that isobutyryl-CoA is a starter unit for isopalmitate biosynthesis and that either acetyl-CoA or butyryl-CoA can be a starter unit for palmitate biosynthesis in S. collinus. However, the pathway for palmitate biosynthesis from acetyl -CoA is less sensitive to thiolactomycin, and it is suggested that the basis for this difference is in the initiation step.