Rapid purification of enzymes of fatty acid biosynthesis from rat adipose tissue.
Acetyl CoA carboxylase, ATP-citrate lyase and fatty acid synthetase were purified to homogeneity by a simple procedure. The purification method consists of polymerization of acetyl CoA carboxylase with citrate followed by avidin-Sepharose affinity chromatography. ATP-citrate lyase and fatty acid synthetase were isolated as by-products of acetyl CoA carboxylase purification and are separated from each other by chromatography on DE-52. ATP-citrate lyase was further purified by CoA-agarose affinity chromatography and fatty acid synthetase was purified on Bio-Gel A-1.5m. Purified ATP-citrate lyase, acetyl CoA carboxylase and fatty acid synthetase had specific activities of 9.9, 2.8 and 1.8 U/mg respectively with an over all recovery of 30, 25 and 50% respectively. Using these purified enzymes, we found that ATP-citrate lyase and acetyl CoA carboxylase were phosphorylated in vitro by both cAMP-dependent protein kinase and ATP-citrate lyase kinase whereas fatty acid synthetase was not phosphorylated by these protein kinases.
D001275:ATP Citrate (pro-S)-Lyase / Q000302:isolation & purification*; D000103:Acetyl-CoA Carboxylase / Q000302:isolation & purification*; D000273:Adipose Tissue / Q000201:enzymology*; D000818:Animals; D004586:Electrophoresis / Q000379:methods; D064429:Fatty Acid Synthases / Q000302:isolation & purification*; D008025:Ligases / Q000302:isolation & purification*; D010766:Phosphorylation; D011494:Protein Kinases / Q000302:isolation & purification; D051381:Rats
D064429:Fatty Acid Synthases; D001275:ATP Citrate (pro-S)-Lyase; D011494:Protein Kinases; D008025:Ligases; D000103:Acetyl-CoA Carboxylase