IDENTITY BETWEEN PALMITOYL-COA SYNTHETASE AND ARACHIDONOYL-COA SYNTHETASE IN HUMAN PLATELETS

Apparent K(m) values have been determined for the substrates ATP, CoA and fatty acids for the long-chain acyl-CoA synthetase (EC 6.2.1.3) reaction in lysates of human blood platelets. The apparent K(m) for ATP was higher for saturated fatty acids (C12:0 to C18:0) than for unsaturated acids (C18:1 to C22:6). Other apparent K(m) values were very similar for all long-chain fatty acids tested. Palmitic acid inhibited the formation of [C-14]arachidonoyl-CoA, and arachidonic acid inhibited the formation of [C-14]palmitoyl-CoA, with [C-14]arachidonate or [C-14]palmitate respectively as substrate. After chromatography of Triton X-100-extracted platelet protein in several systems (hydroxyapatite, DEAE-Sepharose, Sephacryl S-200 HR, CoA-Sepharose, Sephadex G-100 and AcA 34), both arachidonoyl-CoA synthetase and palmitoyl-CoA synthetase activities were eluted together in the various protein peaks, and with approximately the same ratio of activities in all peaks. After some purification steps (DEAE-Sepharose and Sephacryl S-200 HR), the acyl-CoA synthetase activity was up to 37 nmol/min per mg of protein with [C-14]palmitate as substrate, and up to 116 nmol/min per mg of protein with [C-14]arachidonate as substrate. The purification was respectively about 8- and 10-fold. The results indicate that palmitoyl-CoA (or unspecific) synthetase and arachidonoyl-CoA (or specific) synthetase are in fact the same enzyme, in agreement with previously reported results from this laboratory.