1XQL

Effect of a Y265F Mutant on the Transamination Based Cycloserine Inactivation of Alanine Racemase

1XQL の概要

• エントリーDOI: 10.2210/pdb1xql/pdb
• 関連するPDBエントリー: 1SFT 1XQK
• 分子名称: Alanine racemase, 4'-DEOXY-4'-AMINOPYRIDOXAL-5'-PHOSPHATE, (5-HYDROXY-4-{[(3-HYDROXYISOXAZOL-4-YL)AMINO]METHYL}-6-METHYLPYRIDIN-3-YL)METHYL DIHYDROGEN PHOSPHATE, ... (7 entities in total)
• 機能のキーワード: alanine racemase, cycloserine, tim barrel, isomerase
• 由来する生物種: Geobacillus stearothermophilus
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 89335.50

構造登録者

Fenn, T.D.,Holyoak, T.,Stamper, G.F.,Ringe, D. (登録日: 2004-10-12, 公開日: 2005-01-18, 最終更新日: 2023-11-15)

主引用文献

Fenn, T.D.,Holyoak, T.,Stamper, G.F.,Ringe, D.
Effect of a Y265F Mutant on the Transamination-Based Cycloserine Inactivation of Alanine Racemase
Biochemistry, 44:5317-5327, 2005

PubMed Abstract: The requirement for d-alanine in the peptidoglycan layer of bacterial cell walls is fulfilled in part by alanine racemase (EC 5.1.1.1), a pyridoxal 5'-phosphate (PLP)-assisted enzyme. The enzyme utilizes two antiparallel bases focused at the C(alpha) position and oriented perpendicular to the PLP ring to facilitate the equilibration of alanine enantiomers. Understanding how this two-base system is utilized and controlled to yield reaction specificity is therefore a potential means for designing antibiotics. Cycloserine is a known alanine racemase suicide substrate, although its mechanism of inactivation is based on transaminase chemistry. Here we characterize the effects of a Y265F mutant (Tyr265 acts as the catalytic base in the l-isomer case) of Bacillus stearothermophilus alanine racemase on cycloserine inactivation. The Y265F mutant reduces racemization activity 1600-fold [Watanabe, A., Yoshimura, T., Mikami, B., and Esaki, N. (1999) J. Biochem. 126, 781-786] and only leads to formation of the isoxazole end product (the result of the transaminase pathway) in the case of d-cycloserine, in contrast to results obtained using the wild-type enzyme. l-Cycloserine, on the other hand, utilizes a number of alternative pathways in the absence of Y265, emphasizing the importance of Y265 in both the inactivation and racemization pathway. In combination with the kinetics of inactivation, these results suggest roles for each of the two catalytic bases in racemization and inactivation, as well as the importance of Y265 in "steering" the chemistry to favor one pathway over another.

PubMed: 15807525
DOI: 10.1021/bi047842l

実験手法

X-RAY DIFFRACTION (1.8 Å)