Characterization of an arachidonic acid-selective acyl-CoA synthetase from murine T lymphocytes

Abstract

Evidence is presented that the murine thymoma EL4 and cytotoxic T lymphocyte clones possess two distinct long-chain fatty acyl-CoA synthetase activities. One enzyme shows activity toward a broad spectrum of fatty acid substrates, similar to the long-chain fatty acyl-CoA synthetase from rat liver. The other enzyme is selective for arachidonic acid and related fatty acids. Fatty acid competetion studies using EL4 microsomes demonstrate that [ 14C]palmitoyl-CoA synthesis ( K m = 13 ± 1 μM, V max = 7 ± 1 nmol/ mg per min) is inhibited by unlabeled palmitate, oleate, linoleate or linolenate ( K i = 15–25 μM) and weakly by arachidonate ( K i > 100 μM). Similar inhibition is observed for the activation of [ 14C]oleate ( K m = 31 ± 3 μM, V max = 6 ± 2 nmol/mg per min). On the other hand, [ 14C]arachidonyl-CoA synthetase ( K m = 15 ± 3 μM, V max = 13 ± 2 nmol/mg per min) is inhibited by unlabeled arachidonic acid ( k i = 20 μM) but not by unlabeled palmitate, oleate, linoleate and linolenate. The description of arachidonoyl-CoA synthetase in cytotoxic T lymphocyte clones represents the first example of a cell with little or no capacity to synthesize arachidonic acid metabolites, yet which possesses a selective esterification mechanism for the fatty acid. Studies on the specificity of the arachidonic acid-selective acyl-CoA synthetase utilized arachidonic acid metabolites and structurally related fatty acids and yielded two points of interest: (1) metabolism of arachidonic acid to monohydroxy fatty acids (HETEs) resulted in compounds with significantly decreased ability to be activated by the arachidonate-selective acyl-CoA synthetase; (2) arachidonate was a much better substrate than was 5,8,11-eicosatrienoic acid ( K m = 41 μM), the fatty acid which accumulates during essential fatty acid deficiency. The possible role of an arachidonic acid-selective acyl-CoA synthetase in lymphocyte activation and as a homeostatic mechanism during essential fatty acid deficiency is discussed.