Identity between palmitoyl-CoA synthetase and arachidonoyl-CoA synthetase in human platelet?
Apparent Km 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 Km for ATP was higher for saturated fatty acids (C12:0 to C18:0) than for unsaturated acids (C18:1 to C22:6). Other apparent Km values were very similar for all long-chain fatty acids tested. Palmitic acid inhibited the formation of [14C]arachidonoyl-CoA, and arachidonic acid inhibited the formation of [14C]palmitoyl-CoA, with [14C]arachidonate or [14C]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 [14C]palmitate as substrate, and up to 116 nmol/min per mg of protein with [14C]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.
Bakken
A M
AM
Laboratory of Clinical Biochemistry, University of Bergen, Norway.
Farstad
M
M
Holmsen
H
H
D001792:Blood Platelets / Q000201:enzymology*; D002845:Chromatography; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003066:Coenzyme A Ligases / Q000097:blood* / Q000302:isolation & purification; D017886:Durapatite; D006801:Humans; D006882:Hydroxyapatites; D007700:Kinetics; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D013379:Substrate Specificity; D000098859:Long-Chain-Fatty-Acid-CoA Ligase
D006882:Hydroxyapatites; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D017886:Durapatite; D003066:Coenzyme A Ligases; C033896:arachidonate - CoA ligase; C090799:FAA2 protein, S cerevisiae; D000098859:Long-Chain-Fatty-Acid-CoA Ligase