Two distinct long-chain-acyl-CoA synthetases in guinea pig Harderian gland

Two distinct long-chain-acyl-CoA synthetases which have different kinetic properties were identified in the guinea pig Harderian gland. One was localized in the microsomes and the other in the mitochondria. The relative V(max) values of the microsomal enzyme were 8.1, 1.7 and 1 and the apparent Km values were 66.7, 12.0 and 30.0 microM for palmitic, linoleic and arachidonic acids, respectively. The relative V(max) values of the mitochondrial enzyme were 2.7, 3.5 and 1 and the apparent Km values were 33.3, 29.9 and 30.0 microM for palmitic, linoleic and arachidonic acids, respectively. The relative V(max) values for the liver microsomal enzyme were 2.0, 2.5 and 1, while those of the liver mitochondrial enzyme were 4.1, 3.9 and 1 with palmitic, linoleic and arachidonic acids, respectively. There were no difference between the microsomal and the mitochondrial enzymes in the liver, regarding apparent Km values; these were 38.4, 29.9 and 22.0 microM for palmitic, linoleic and arachidonic acids, respectively. Thus, the substrate specificity and catalytic rate of the mitochondrial enzyme in Harderian gland for palmitic, linoleic and arachidonic acids were similar to the liver enzyme, but not to the microsomal enzyme in Harderian gland. On the other hand, the antiserum raised against the rat liver enzyme immune-titrated and immuno-blotted the enzymes from Harderian gland microsomes and liver, but not so the enzyme from Harderian gland mitochondria. Thus, the microsomal enzyme in Harderian gland had a common immunogenic epitope(s) with the liver enzyme, but the mitochondrial enzyme did not. The Harderian gland mitochondrial enzyme was a distinct protein from liver enzymes. The catalytic and immunogenic characteristics suggest that the enzyme proteins in the Harderian gland are unique, that is, different from that in the liver. The large V(max) value of the Harderian gland microsomal enzyme for palmitic acid suggests that it contributes to the synthesis of a large amount of the secretory lipid and the high Km value to maintenance of cellular lipid in this organ. The evidence that long-chain-acyl-CoA synthetase in the mitochondria is distinct from that in the microsomes was first found in guinea pig Harderian gland.