1GES

ANATOMY OF AN ENGINEERED NAD-BINDING SITE

1GES の概要

• エントリーDOI: 10.2210/pdb1ges/pdb
• 分子名称: GLUTATHIONE REDUCTASE, FLAVIN-ADENINE DINUCLEOTIDE (3 entities in total)
• 機能のキーワード: oxidoreductase(flavoenzyme)
• 由来する生物種: Escherichia coli
• 細胞内の位置: Cytoplasm: P06715
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 99049.36

構造登録者

Mittl, P.R.E.,Schulz, G.E. (登録日: 1994-01-18, 公開日: 1994-11-01, 最終更新日: 2024-10-09)

主引用文献

Mittl, P.R.,Berry, A.,Scrutton, N.S.,Perham, R.N.,Schulz, G.E.
Anatomy of an engineered NAD-binding site.
Protein Sci., 3:1504-1514, 1994

PubMed Abstract: The coenzyme specificity of Escherichia coli glutathione reductase was switched from NADP to NAD by modifying the environment of the 2'-phosphate binding site through a set of point mutations: A179G, A183G, V197E, R198M, K199F, H200D, and R204P (Scrutton NS, Berry A, Perham RN, 1990, Nature 343:38-43). In order to analyze the structural changes involved, we have determined 4 high-resolution crystal structures, i.e., the structures of the wild-type enzyme (1.86 A resolution, R-factor of 16.8%), of the wild-type enzyme ligated with NADP (2.0 A, 20.8%), of the NAD-dependent mutant (1.74 A, 16.8%), and of the NAD-dependent mutant ligated with NAD (2.2 A, 16.9%). A comparison of these structures reveals subtle differences that explain details of the specificity change. In particular, a peptide rotation occurs close to the adenosine ribose, with a concomitant change of the ribose pucker. The mutations cause a contraction of the local chain fold. Furthermore, the engineered NAD-binding site assumes a less rigid structure than the NADP site of the wild-type enzyme. A superposition of the ligated structures shows a displacement of NAD versus NADP such that the electron pathway from the nicotinamide ring to FAD is elongated, which may explain the lower catalytic efficiency of the mutant. Because the nicotinamide is as much as 15 A from the sites of the mutations, this observation reminds us that mutations may have important long-range consequences that are difficult to anticipate.

PubMed: 7833810

実験手法

X-RAY DIFFRACTION (1.74 Å)