title: Cloning, purification, crystallization and preliminary X-ray crystallographic analysis of the biosynthetic N-acetylornithine aminotransferases from Salmonella typhimurium and Escherichia coli.
abstract: Acetylornithine aminotransferase (AcOAT) is a type I pyridoxal 5'-phosphate-dependent enzyme catalyzing the conversion of N-acetylglutamic semialdehyde to N-acetylornithine in the presence of alpha-ketoglutarate, a step involved in arginine metabolism. In Escherichia coli, the biosynthetic AcOAT also catalyzes the conversion of N-succinyl-L-2-amino-6-oxopimelate to N-succinyl-L,L-diaminopimelate, one of the steps in lysine biosynthesis. It is closely related to ornithine aminotransferase. AcOAT was cloned from Salmonella typhimurium and E. coli, overexpressed in E. coli and purified using Ni-NTA affinity column chromatography. The enzymes crystallized in the presence of gabaculine. Crystals of E. coli AcOAT (eAcOAT) only diffracted X-rays to 3.5 A and were twinned. The crystals of S. typhimurium AcOAT (sAcOAT) diffracted to 1.9 A and had a dimer in the asymmetric unit. The structure of sAcOAT was solved by the molecular-replacement method.
authors.affiliation: Molecular Biophysics Unit, Indian Institute of Science, Bangalore 560 012, India.
authors.forename: V
authors.initials: V
authors.lastname: Rajaram
authors.forename: K
authors.initials: K
authors.lastname: Prasad
authors.forename: P
authors.initials: P
authors.lastname: Ratna Prasuna
authors.forename: N
authors.initials: N
authors.lastname: Ramachandra
authors.forename: S R
authors.initials: SR
authors.lastname: Bharath
authors.forename: H S
authors.initials: HS
authors.lastname: Savithri
authors.forename: M R N
authors.initials: MR
authors.lastname: Murthy
mesh_terms: D003001:Cloning, Molecular; D003460:Crystallization; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000201:enzymology*; D012486:Salmonella typhimurium / Q000201:enzymology*; D000637:Transaminases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification
chemical_list: D000637:Transaminases; C018731:acetylornithine transaminase