Effect of phospholipase A2 and free fatty acids on lipid-protein interactions in long- and very-long-chain fatty acyl-CoA elongation enzyme systems of brain microsomes.
The elucidation of the mechanism of phospholipase A2-induced inactivation of the condensation enzyme provided evidence concerning the important role of lipid-enzyme interactions in maintaining the condensation activity in swine cerebral microsomes. A quantitative analysis of fatty acid release by phospholipase A2 from the microsomal membrane revealed that only 5 nmol of free fatty acid per mg microsomal protein was released, including oleic acid and arachidonic acid, by treatment with 0.4 unit of phospholipase A2 per mg microsomal protein for 15 s at 23 degrees C. Under these conditions, the condensation activity for endogenous 16:0-CoA and 20:4-CoA decreased to half and that for exogenous 20:0-CoA decreased to 75%. However, the addition of free fatty acids and lysophospholipids or a mixture of them at 5-10 nmol/mg protein did not change the condensation activity for endogenous 16:0-CoA and 20:4-CoA, or for exogenous 20:0-CoA. These results indicated that phospholipase A2 inhibited the condensation activity by acting directly on phospholipids that are indispensable to maintaining the function of the condensation enzyme. The Arrhenius plot for the condensation of endogenous 16:0-CoA showed a break at around 16 degrees C, whereas no break of the plot was observed for the condensation of 20:0-CoA and 20:4-CoA. The activation energy for the condensation of 16:0-CoA and 20:4-CoA was decreased by the addition of free fatty acids such as oleic acid and stearic acid, with disappearance of the Arrhenius break for 16:0-CoA condensation, whereas the activation energy for the condensation of 20:0-CoA was not changed. These results suggest that the type of lipid-protein interaction in the condensation enzyme for 20:0-CoA is different from that for 16:0-CoA and 20:4-CoA.
Yoshida
S
S
Research Laboratory Center, Medical College of Oita, Japan.
Saitoh
T
T
Takeshita
M
M
D015099:3-Oxoacyl-(Acyl-Carrier-Protein) Synthase / Q000378:metabolism*; D000214:Acyl Coenzyme A / Q000378:metabolism*; D000217:Acyltransferases / Q000378:metabolism*; D000818:Animals; D016718:Arachidonic Acid; D001095:Arachidonic Acids / Q000378:metabolism; D001921:Brain / Q000201:enzymology* / Q000648:ultrastructure; D003066:Coenzyme A Ligases / Q000378:metabolism; D005227:Fatty Acids / Q000378:metabolism; D005230:Fatty Acids, Nonesterified / Q000494:pharmacology*; D008246:Lysophospholipids / Q000378:metabolism / Q000494:pharmacology; D008861:Microsomes / Q000187:drug effects / Q000201:enzymology; D019301:Oleic Acid; D009829:Oleic Acids / Q000494:pharmacology; D010740:Phospholipases / Q000494:pharmacology*; D010741:Phospholipases A / Q000494:pharmacology*; D054467:Phospholipases A2; D010743:Phospholipids / Q000378:metabolism*; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D013229:Stearic Acids / Q000494:pharmacology; D013552:Swine; D013816:Thermodynamics; D000098859:Long-Chain-Fatty-Acid-CoA Ligase
D000214:Acyl Coenzyme A; D001095:Arachidonic Acids; D005227:Fatty Acids; D005230:Fatty Acids, Nonesterified; D008246:Lysophospholipids; D009829:Oleic Acids; D010743:Phospholipids; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D013229:Stearic Acids; D016718:Arachidonic Acid; D019301:Oleic Acid; C031183:stearic acid; D000217:Acyltransferases; D015099:3-Oxoacyl-(Acyl-Carrier-Protein) Synthase; D010740:Phospholipases; D010741:Phospholipases A; D054467:Phospholipases A2; D003066:Coenzyme A Ligases; C090799:FAA2 protein, S cerevisiae; D000098859:Long-Chain-Fatty-Acid-CoA Ligase