Identity of Long-Chain Acyl-Coenzyme A Synthetase of Microsomes, Mitochondria, and Peroxisomes in Rat Liver1

Abstract

The identity of long-chain acyl-CoA synthetase in microsomes, mitochondria, and peroxisomes of rat liver was examined by using the antibody raised against a purified preparation of the microsomal enzyme. The enzyme activities of these three organelles and the purified microsomal enzyme were titrated by the antibody in a very similar fashion when the activity was measured in terms of palmitoyl-CoA synthetase activity. It was shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the immunoprecipitates and by Western blot analysis that the enzymes of all three organelles consisted of a polypeptide with the same molecular weight as that of the purified enzyme, and that the specific enzyme activity of the antigenic protein in all three subcellular compartments was nearly the same. The presence of other palmitoyl-CoA synthetase activity in these organelles could not be confirmed. Immunocytochemical study to locate the antigenic site with protein A-gold complex showed that the gold particles were closely associated with the membranes of these organelles. The cell-free translation product in a rabbit reticulocyte lysate protein-synthesizing system and the subunit of the mature enzyme labeled with [35S]methionine in the liver slices exhibited the same mobility as the subunit of the purified enzyme on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The enzyme in microsomes, mitochondria, and peroxisomes was labeled at nearly the same rate when the liver slices were incubated with [35S]methionine.

Peroxisomal fatty acid beta-oxidation in relation to the accumulation of very long chain fatty acids in cultured skin fibroblasts from patients with Zellweger syndrome and other peroxisomal disorders.

“…The results of Fig. 3 show that palmitoyl-CoA synthetase is present in peroxisomes, mitochondria and microsomes, in agreement with data from a number of authors (16)(17)(18)(19). In contrast, however, very long chain fatty acyl-CoA synthetase showed a bimodal distribution of activity; the enzyme is present only in peroxisomes and microsomes and not in mitochondria.…”

“…Several acyl-CoA synthetases are known to be present in mammalian cells (14,15) including long chain acyl-CoA synthetase (acid:CoA ligase (AMP-forming), EC 6.2.1.3; also known as palmitoyl-CoA synthetase), which has been found to be localized in mitochondria and microsomes (for reviews see 14,15) as well as in peroxisomes (16)(17)(18)(19). Evidence has recently been brought forward (20) (25) were measured according to published procedures.…”

Enzymatic and Genetic Characterization of Firefly Luciferase andDrosophilaCG6178 as a Fatty Acyl-CoA Synthetase

“…The efficient substrates for rat Acsl3 are lauric acid (C12:0), myristic acid (C14:0), arachidonic acid (C20:4), and EPA (C20:5). 19) Furthermore, rat ACSL activity was observed in peroxisomes, 22,23) as in firefly luciferase. 24) Firefly luciferases and CG6178 possess a peroxisomal targeting signal sequence at the C-terminus.…”

Peroxisomal and Mitochondrial Fatty Acid β-Oxidation in Mice Nullizygous for Both Peroxisome Proliferator-activated Receptor α and Peroxisomal Fatty Acyl-CoA Oxidase

“…The content of hepatic VLCAS was not much different among AOX Ϫ/Ϫ , PPAR␣ Ϫ/Ϫ , and PPAR␣ Ϫ/Ϫ AOX Ϫ/Ϫ mice ( Table I). The amount of PCS, an enzyme with tripartite (microsomal, mitochondrial, and peroxisomal) distribution (20), increased ϳ2-fold in wild type mice treated with ciprofibrate (Table I). The hepatic PCS content in AOX Ϫ/Ϫ mice maintained on control diet was also elevated with no further increase occurring when these mice were treated with ciprofibrate (Table I).…”