Fatty acids bound to unilamellar lipid vesicles as substrates for microsomal acyl-CoA ligase.
Palmitate incorporated into single-layered vesicles of phosphatidylcholine was used as a substrate for palmitoyl coenzyme A ligase (palmitoyl-CoA ligase) in microsomes from rat liver. This was done in order to avoid the use of detergents for dispersal of the water-insoluble palmitate and the possibility of precipitating palmitate added to the aqueous assay as a salt suspension. The activity of the ligase measured when palmitate was added to assays as a component of phospholipid vesicles was 10-40-fold greater vs. activities reported in the literature using other methods for adding fatty acids to the assay system. Phospholipids, however, had no direct effect on the activity of palmitoyl-CoA ligase. The data indicate, therefore, that the activity of this enzyme has been underestimated because of the manner in which fatty acid was added to the assay, which has a significant effect on the activity of the ligase. It is shown too that the rate of spontaneous transfer of palmitate from unilamellar vesicles of phosphatidylcholine to microsomes via a hydrated intermediate is far more rapid than the inherent catalytic activity of the fatty acyl-CoA ligase. The data also suggest that the membrane-associated pool of fatty acid and not fatty acid in the aqueous phase of the assay is the pool of substrate interacting with the ligase.
Noy
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Zakim
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D000818:Animals; D002250:Carbon Radioisotopes; D002352:Carrier Proteins / Q000378:metabolism; D003066:Coenzyme A Ligases / Q000378:metabolism*; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D007413:Intestinal Mucosa / Q000378:metabolism; D007700:Kinetics; D008099:Liver / Q000378:metabolism; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology*; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D019308:Palmitic Acid; D010169:Palmitic Acids / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D012709:Serum Albumin / Q000378:metabolism; D013229:Stearic Acids / Q000378:metabolism; D013379:Substrate Specificity; D000098859:Long-Chain-Fatty-Acid-CoA Ligase
D002250:Carbon Radioisotopes; D002352:Carrier Proteins; C490174:Fabp7 protein, rat; D000071438:Fatty Acid-Binding Protein 7; D050556:Fatty Acid-Binding Proteins; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D010169:Palmitic Acids; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D012709:Serum Albumin; D013229:Stearic Acids; D019308:Palmitic Acid; C031183:stearic acid; D003066:Coenzyme A Ligases; C090799:FAA2 protein, S cerevisiae; D000098859:Long-Chain-Fatty-Acid-CoA Ligase