title: Purification and characterization of ornithine acetyltransferase from Saccharomyces cerevisiae.
abstract: Ornithine acetyltransferase has been purified 4000-fold to homogeneity from Saccharomyces cerevisiae. The enzyme catalyses the freely reversible interchange of an acetyl group between N-acetylornithine and glutamate and has a specific activity of 22 mumol.min-1.mg-1 at 37 degrees C and pH 7.5. The Km values were determined for the substrates of the forward and reverse directions to be 1.0 mM for N-acetylornithine, 7.2 mM for glutamate, 1.5 mM for ornithine and 17.1 mM for N-acetylglutamate. The enzyme was localised to the mitochondrial matrix and was found to be a 57-kDa heterodimer consisting of subunits of 31 kDa and 26 kDa. Antibodies raised against the small subunit immunoprecipitated a single in vitro translation product of approximately 57 kDa suggesting that the subunits are processed from a single precursor protein. This is supported by N-terminal sequence analysis which shows that the 26-kDa subunit exhibits 40% sequence identity (8 out of 20) with the N-terminus of ornithine acetyltransferase from Neisseria gonorrhoeae whereas the N-terminus of the 31-kDa subunit exhibits 45% identity (9 out of 20) with a sequence located in the middle of the 60-kDa N. gonorrhoeae enzyme. The N-terminal sequence of the small subunit has the potential to form an amphiphilic helix, further suggesting that the precursor protein with the small subunit at its N-terminus could be targeted to mitochondria and processed into two subunits.
authors.affiliation: School of Biochemistry, La Trobe University, Bundoora, Australia.
authors.forename: Y
authors.initials: Y
authors.lastname: Liu
authors.forename: R
authors.initials: R
authors.lastname: Van Heeswijck
authors.forename: P
authors.initials: P
authors.lastname: Høj
authors.forename: N
authors.initials: N
authors.lastname: Hoogenraad
mesh_terms: D000123:Acetyltransferases / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification*; D000595:Amino Acid Sequence; D000818:Animals; D007700:Kinetics; D008969:Molecular Sequence Data; D011817:Rabbits; D012441:Saccharomyces cerevisiae / Q000201:enzymology*
chemical_list: D000123:Acetyltransferases; C020646:glutamate acetyltransferase