Subunits of fatty acid synthetase complexes. Enzymatic activities and properties of the half-molecular weight nonidentical subunits of pigeon liver fatty acid synthetase.
The separation of the half-molecular weight, nonidentical subunits (I and II) of the pigeon liver fatty acid synthetase complex has been achieved on a large (20 mg) scale by affinity chromatography on Sepharose epsilon-aminocaproyl pantetheine. This separation requires a careful control of temperature, ionic strength, pH, and column flow rate for success. The yield of subunit II is further improved by transacetylation (with acetyl-CoA) of the dissociated fatty acid synthetase prior to affinity chromatography. The separated subunit I (reductase) contains the 4'-phosphopantetheine (A2) acyl binding site, two NADPH binding sites, and beta-ketoacyl and crotonyl thioester reductases. Subunit II (transacylase) contains the B1 (hydroxyl or loading) and B2 (cysteine) acyl binding sites, and acetyl- and malonyl-CoA: pantetheine transacylases. When subunit I is mixed in equimolar quantities with subunit II, an additional NADPH binding site is found even though subunit II alone shows no NADPH binding. Both subunits contain activities for the partial reactions, beta-hydroxybutyryl thioester dehydrase (crotonase) and palmityl-CoA deacylase. Subunit I has 8 sulfhydryl groups per mol whereas subunit II has 60. Reconstitution of fatty acid synthetase activity to 75% of the control level is achieved on reassociation of subunits I and II.
D000123:Acetyltransferases / Q000378:metabolism; D000217:Acyltransferases / Q000378:metabolism; D000818:Animals; D002499:Centrifugation, Density Gradient; D002846:Chromatography, Affinity; D003065:Coenzyme A / Q000378:metabolism; D010856:Columbidae; D004591:Electrophoresis, Polyacrylamide Gel; D064429:Fatty Acid Synthases / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D046911:Macromolecular Substances; D008314:Malonates; D008970:Molecular Weight; D009249:NADP / Q000494:pharmacology; D010084:Oxidation-Reduction; D013997:Time Factors
D046911:Macromolecular Substances; D008314:Malonates; D009249:NADP; D000217:Acyltransferases; D000123:Acetyltransferases; D064429:Fatty Acid Synthases; D003065:Coenzyme A