Purification and properties of rat liver peroxisomal very-long-chain acyl-CoA synthetase.
It is generally accepted that there are two different acyl-CoA synthetases in rat liver peroxisomes. One is long-chain acyl-CoA synthetase, and the other very-long-chain acyl-CoA synthetase. Nowadays, the nature of long-chain acyl-CoA synthetase is well-known, but that of very-long-chain acyl-CoA synthetase remains unclear. Very-long-chain acyl-CoA synthetase has been extracted from the washed membrane fraction of frozen rat liver peroxisomes with a buffer containing a detergent, and has been purified by chromatography on Ultrogel AcA 34, calcium phosphate gel/cellulose, blue dextran-Sepharose 4B and DEAE-Toyopearl. The molecular masses of the native enzyme and the subunit were estimated to be 235 and 70 kDa, respectively. This enzyme showed marked differences in behavior from long-chain acyl-CoA synthetase during purification. The carbon chain length specificity of very-long-chain acyl-CoA synthetase differed from that of long-chain acyl-CoA synthetase. Very-long-chain acyl-CoA synthetase was active toward long- and very-long-chain fatty acids, but more active toward very-long-chain fatty acids compared with long-chain acyl-CoA synthetase. Antibodies against long-chain acyl-CoA synthetase did not cross-react to very-long-chain acyl-CoA synthetase. Based on these data, the final enzyme preparation is judged to be highly purified very-long-chain acyl-CoA synthetase from rat liver peroxisomes.
Uchida
Y
Y
Department of Pediatrics, Gifu University School of Medicine.
Kondo
N
N
Orii
T
T
Hashimoto
T
T
D000214:Acyl Coenzyme A / Q000378:metabolism; D044942:Acyl-CoA Dehydrogenase, Long-Chain / Q000302:isolation & purification* / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D003902:Detergents / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008099:Liver / Q000201:enzymology* / Q000648:ultrastructure; D008297:Male; D008830:Microbodies / Q000201:enzymology*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D051381:Rats; D017208:Rats, Wistar; D013379:Substrate Specificity
D000214:Acyl Coenzyme A; D003902:Detergents; C018582:lignoceroyl-coenzyme A; D000255:Adenosine Triphosphate; D044942:Acyl-CoA Dehydrogenase, Long-Chain