Article (Scientific journals)
Glycoside hydrolase family 5: structural snapshots highlighting the involvement of two conserved residues in catalysis
Collet, Laetitia; Vander Wauven, Corinne; Dutoit, Raphaël et al.
2021In Acta Crystallographica. Section D, Structural Biology
Peer Reviewed verified by ORBi
 

Files


Full Text
actaD_manuscript_reviewed.docx
Author postprint (4.87 MB)
Request a copy

All documents in ORBi are protected by a user license.

Send to



Details



Abstract :
[en] 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.
Research center :
Institut de recherches LABIRIS
Disciplines :
Biochemistry, biophysics & molecular biology
Author, co-author :
Collet, Laetitia ;  Université de Liège - ULiège > Form. doct. sc. (bioch., biol. mol. cel., bioinf. - paysage)
Vander Wauven, Corinne
Dutoit, Raphaël
Oudjama, Yamina
Galleni, Moreno ;  Université de Liège - ULiège > Département des sciences de la vie > Macromolécules biologiques
Language :
English
Title :
Glycoside hydrolase family 5: structural snapshots highlighting the involvement of two conserved residues in catalysis
Publication date :
08 February 2021
Journal title :
Acta Crystallographica. Section D, Structural Biology
eISSN :
2059-7983
Publisher :
John Wiley and Sons, Hoboken, United States - New Jersey
Peer reviewed :
Peer Reviewed verified by ORBi
Name of the research project :
Caractérisation de RBcel1: une endoglucanase dotée d’activité transglycosylase
Funders :
COCOF - Commission Communautaire Française [BE]
Available on ORBi :
since 09 March 2021

Statistics


Number of views
47 (11 by ULiège)
Number of downloads
1 (1 by ULiège)

Scopus citations®
 
3
Scopus citations®
without self-citations
2
OpenCitations
 
2

Bibliography


Similar publications



Contact ORBi