5LJF

Crystal structure of the endo-1,4-glucanase RBcel1 E135A with cellotriose

5LJF の概要

• エントリーDOI: 10.2210/pdb5ljf/pdb
• 関連するPDBエントリー: 4EE9 4M24
• 関連するBIRD辞書のPRD_ID: PRD_900021
• 分子名称: Endoglucanase, beta-D-glucopyranose-(1-4)-beta-D-glucopyranose-(1-4)-beta-D-glucopyranose (3 entities in total)
• 機能のキーワード: glycosyl hydrolase family 5, cellulase, tim barrel, beta-1, 4-endoglucanase, hydrolase
• 由来する生物種: uncultured bacterium
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 73656.91

構造登録者

Dutoit, R.,Collet, L.,Galleni, M.,Bauvois, C. (登録日: 2016-07-18, 公開日: 2017-08-02, 最終更新日: 2024-10-16)

主引用文献

Collet, L.,Vander Wauven, C.,Oudjama, Y.,Galleni, M.,Dutoit, R.
Glycoside hydrolase family 5: structural snapshots highlighting the involvement of two conserved residues in catalysis.
Acta Crystallogr D Struct Biol, 77:205-216, 2021

PubMed Abstract: The ability of retaining glycoside hydrolases (GHs) to transglycosylate is inherent to the double-displacement mechanism. Studying reaction intermediates, such as the glycosyl-enzyme intermediate (GEI) and the Michaelis complex, could provide valuable information to better understand the molecular factors governing the catalytic mechanism. Here, the GEI structure of RBcel1, an endo-1,4-β-glucanase of the GH5 family endowed with transglycosylase activity, is reported. It is the first structure of a GH5 enzyme covalently bound to a natural oligosaccharide with the two catalytic glutamate residues present. The structure of the variant RBcel1_E135A in complex with cellotriose is also reported, allowing a description of the entire binding cleft of RBcel1. Taken together, the structures deliver different snapshots of the double-displacement mechanism. The structural analysis revealed a significant movement of the nucleophilic glutamate residue during the reaction. Enzymatic assays indicated that, as expected, the acid/base glutamate residue is crucial for the glycosylation step and partly contributes to deglycosylation. Moreover, a conserved tyrosine residue in the -1 subsite, Tyr201, plays a determinant role in both the glycosylation and deglycosylation steps, since the GEI was trapped in the RBcel1_Y201F variant. The approach used to obtain the GEI presented here could easily be transposed to other retaining GHs in clan GH-A.

PubMed: 33559609
DOI: 10.1107/S2059798320015557

実験手法

X-RAY DIFFRACTION (1.73439601224 Å)