2byl
STRUCTURE OF ORNITHINE AMINOTRANSFERASE TRIPLE MUTANT Y85I Y55A G320F
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OverviewOverview
Ornithine aminotransferase and 4-aminobutyrate aminotransferase are, related pyridoxal phosphate-dependent enzymes having different substrate, specificities. The atomic structures of these enzymes have shown (i) that, active site differences are limited to the steric positions occupied by, two tyrosine residues in ornithine aminotransferase and (ii) that, uniquely among related, structurally characterized aminotransferases, the, conserved arginine that binds the alpha-carboxylate of alpha-amino acids, interacts tightly with a glutamate residue. To determine the contribution, of these residues to the specificities of the enzymes, we analyzed, site-directed mutants of ornithine aminotransferase by rapid reaction, kinetics, x-ray crystallography, and 13C NMR spectroscopy. Mutation of one, tyrosine (Tyr-85) to isoleucine, as found in aminobutyrate, aminotransferase, decreased the rate of the reaction of the enzyme with, ornithine 1000-fold and increased that with 4-aminobutyrate 16-fold, indicating that Tyr-85 is a major determinant of specificity toward, ornithine. Unexpectedly, the limiting rate of the second half of the, reaction, conversion of ketoglutarate to glutamate, was greatly increased, although the kinetics of the reverse reaction were unaffected. A mutant in, which the glutamate (Glu-235) that interacts with the conserved arginine, was replaced by alanine retained its regiospecificity for the delta-amino, group of ornithine, but the glutamate reaction was enhanced 650-fold, whereas only a 5-fold enhancement of the ketoglutarate reaction rate, resulted. A model is proposed in which conversion of the enzyme to its, pyridoxamine phosphate form disrupts the internal glutamate-arginine, interaction, thus enabling ketoglutarate but not glutamate to be a good, substrate.
DiseaseDisease
Known disease associated with this structure: Gyrate atrophy of choroid and retina with ornithinemia, B6 responsive or unresponsive OMIM:[258870]
About this StructureAbout this Structure
2BYL is a Single protein structure of sequence from Homo sapiens with PLP as ligand. Active as Ornithine aminotransferase, with EC number 2.6.1.13 Structure known Active Site: AC1. Full crystallographic information is available from OCA.
ReferenceReference
Determinants of substrate specificity in omega-aminotransferases., Markova M, Peneff C, Hewlins MJ, Schirmer T, John RA, J Biol Chem. 2005 Oct 28;280(43):36409-16. Epub 2005 Aug 11. PMID:16096275
Page seeded by OCA on Mon Nov 12 21:08:13 2007