2BYL

Structure of ornithine aminotransferase triple mutant Y85I Y55A G320F

2BYL の概要

• エントリーDOI: 10.2210/pdb2byl/pdb
• 関連するPDBエントリー: 1GBN 1OAT 2BYJ 2CAN 2OAT
• 分子名称: ORNITHINE AMINOTRANSFERASE, PYRIDOXAL-5'-PHOSPHATE (3 entities in total)
• 機能のキーワード: transferase, disease mutation, mitochondrion, transit peptide plp-dependent enzyme, polymorphism, pyridoxal phosphate
• 由来する生物種: HOMO SAPIENS (HUMAN)
• 細胞内の位置: Mitochondrion matrix: P04181
• タンパク質・核酸の鎖数: 3
• 化学式量合計: 146366.45

構造登録者

Markova, M.,Peneff, C.,Hewlins, M.J.E.,Schirmer, T.,John, R.A. (登録日: 2005-08-03, 公開日: 2005-09-07, 最終更新日: 2023-12-13)

主引用文献

Markova, M.,Peneff, C.,Hewlins, M.J.E.,Schirmer, T.,John, R.A.
Determinants of Substrate Specificity in Omega-Aminotransferases.
J.Biol.Chem., 280:36409-, 2005

PubMed Abstract: 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.

PubMed: 16096275
DOI: 10.1074/JBC.M506977200

実験手法

X-RAY DIFFRACTION (2.15 Å)