Purification and properties of carnitine acetyltransferase from rat liver.
Carnitine acetyltransferase was purified from rat liver after induction of the enzyme by feeding with di(2-ethylhexyl)phthalate. Two enzyme sources were used: the mitochondrial fraction and the homogenate of the liver. The purification procedure was essentially the same for the two enzyme sources. The enzyme purified from the mitochondrial fraction consisted of two different polypeptides with molecular weights of 36,500 and 27,000, whereas that from the homogenate consisted of one polypeptide with a molecular weight of 67,500. Amino acid compositions and peptide maps of the limited proteolytic products of the two enzyme preparations were nearly the same. Their antibodies were cross-reactive. Catalytic properties of the two preparations were nearly the same: the specific enzyme activities, double reciprocal plots of initial velocity study, substrate specificities for acylcarnitines having various carbon chain lengths, apparent Michaelis constants for substrates. On electrophoresis of the immunoprecipitate obtained after incubation of the mitochondrial extract, the two immunoreactive polypeptides with molecular weights of 36,500 and 27,000 were found. But only one polypeptide, with molecular weight of 67,500, was detected when the protease inhibitors were added to the mitochondrial extract. It was concluded that the enzyme in the mitochondrial fraction was a monomeric form but was converted into a dimeric form by proteolytic modification after the disruption of mitochondria. The preparation from the post-mitochondrial fraction, which had a lower specific activity, contained two polypeptides whose molecular weights were 69,000 and 67,500. They could not be separated from each other throughout the purification. The peptide maps of the products of the limited proteolysis were very similar.
D000123:Acetyltransferases / Q000302:isolation & purification*; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002332:Carnitine O-Acetyltransferase / Q000302:isolation & purification* / Q000378:metabolism; D005779:Immunodiffusion; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D046911:Macromolecular Substances; D008297:Male; D008970:Molecular Weight; D051381:Rats; D011919:Rats, Inbred Strains; D013379:Substrate Specificity
D000596:Amino Acids; D046911:Macromolecular Substances; D000123:Acetyltransferases; D002332:Carnitine O-Acetyltransferase