title: In vivo and in vitro effects of thiolactomycin on fatty acid biosynthesis in Streptomyces collinus.
abstract: A stable-isotope assay was used to analyze the effectiveness of various perdeuterated short-chain acyl coenzyme A (acyl-CoA) compounds as starter units for straight- and branched-chain fatty acid biosynthesis in cell extracts of Streptomyces collinus. In these extracts perdeuterated isobutyryl-CoA was converted to isopalmitate (a branched-chain fatty acid), while butyryl-CoA was converted to palmitate (a straight-chain fatty acid). These observations are consistent with previous in vivo analyses of fatty acid biosynthesis in S. collinus, which suggested that butyryl-CoA and isobutyryl-CoA function as starter units for palmitate and isopalmitate biosynthesis, respectively. Additionally, in vitro analysis demonstrated that acetyl-CoA can function as a starter unit for palmitate biosynthesis. Palmitate biosynthesis and isopalmitate biosynthesis in these cell extracts were both effectively inhibited by thiolactomycin, a known type II fatty acid synthase inhibitor. In vivo experiments demonstrated that concentrations of thiolactomycin ranging from 0.1 to 0.2 mg/ml produced both a dramatic decrease in the cellular levels of branched-chain fatty acids and a surprising three- to fivefold increase in the cellular levels of the straight-chain fatty acids palmitate and myristate. Additional in vivo incorporation studies with perdeuterated butyrate suggested that, in accord with the in vitro studies, the biosynthesis of the palmitate from butyryl-CoA decreases in the presence of thiolactomycin. In contrast, in vivo incorporation studies with perdeuterated acetate demonstrated that the biosynthesis of palmitate from acetyl-CoA increases in the presence of thiolactomycin. 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.
authors.affiliation: Department of Pharmaceutical Sciences, School of Pharmacy, University of Maryland at Baltimore, 21201, USA.
authors.forename: K K
authors.initials: KK
authors.lastname: Wallace
authors.forename: S
authors.initials: S
authors.lastname: Lobo
authors.forename: L
authors.initials: L
authors.lastname: Han
authors.forename: H A
authors.initials: HA
authors.lastname: McArthur
authors.forename: K A
authors.initials: KA
authors.lastname: Reynolds
mesh_terms: D019342:Acetic Acid / Q000378:metabolism; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D002087:Butyrates / Q000378:metabolism; D020148:Butyric Acid; D064429:Fatty Acid Synthases / Q000378:metabolism; D005227:Fatty Acids / Q000096:biosynthesis*; D058610:Isobutyrates; D019308:Palmitic Acid / Q000378:metabolism; D013302:Streptomyces / Q000378:metabolism*; D013876:Thiophenes / Q000494:pharmacology
chemical_list: D000900:Anti-Bacterial Agents; D002087:Butyrates; D005227:Fatty Acids; D058610:Isobutyrates; D013876:Thiophenes; D020148:Butyric Acid; D019308:Palmitic Acid; C035068:thiolactomycin; C020380:isobutyric acid; D064429:Fatty Acid Synthases; D019342:Acetic Acid