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J Biol Chem. 2016 Mar 25;291(13):7183-94. doi: 10.1074/jbc.M115.695999. Epub 2016 Jan 11.

The Quaternary Structure of a Glycoside Hydrolase Dictates Specificity toward β-Glucans.

Author information

1
From the Institut des Sciences Moléculaires de Marseille-BiosCiences, UMR7313 CNRS, Aix-Marseille University, Pôle de l'Etoile, 13284 Marseille, France, the INRA, UMR1163, Biodiversité et Biotechnologie Fongiques, Aix-Marseille University, Polytech'Marseille, F-13288 Marseille, France.
2
the Architecture et Fonction des Macromolécules Biologiques, UMR7257 CNRS, Aix-Marseille University, F-13288 Marseille, France, the INRA, USC1408 Architecture et Fonction des Macromolécules Biologiques, F-13288 Marseille, France, and.
3
the Architecture et Fonction des Macromolécules Biologiques, UMR7257 CNRS, Aix-Marseille University, F-13288 Marseille, France.
4
the Architecture et Fonction des Macromolécules Biologiques, UMR7257 CNRS, Aix-Marseille University, F-13288 Marseille, France, the INRA, USC1408 Architecture et Fonction des Macromolécules Biologiques, F-13288 Marseille, France, and the Department of Biological Sciences, King Abdulaziz University, Jeddah 21589, Saudi Arabia.
5
the INRA, UMR1163, Biodiversité et Biotechnologie Fongiques, Aix-Marseille University, Polytech'Marseille, F-13288 Marseille, France, jean-guy.berrin@univ-amu.fr.

Abstract

In the Carbohydrate-Active Enzyme (CAZy) database, glycoside hydrolase family 5 (GH5) is a large family with more than 6,000 sequences. Among the 51 described GH5 subfamilies, subfamily GH5_26 contains members that display either endo-β(1,4)-glucanase or β(1,3;1,4)-glucanase activities. In this study, we focused on the GH5_26 enzyme fromSaccharophagus degradans(SdGluc5_26A), a marine bacterium known for its capacity to degrade a wide diversity of complex polysaccharides.SdGluc5_26A displays lichenase activity toward β(1,3;1,4)-glucans with a side cellobiohydrolase activity toward β(1,4)-glucans. The three-dimensional structure ofSdGluc5_26A adopts a stable trimeric quaternary structure also observable in solution. The N-terminal region ofSdGluc5_26A protrudes into the active site of an adjacent monomer. To understand whether this occupation of the active site could influence its activity, we conducted a comprehensive enzymatic characterization ofSdGluc5_26A and of a mutant truncated at the N terminus. Ligand complex structures and kinetic analyses reveal that the N terminus governs the substrate specificity ofSdGluc5_26A. Its deletion opens the enzyme cleft at the -3 subsite and turns the enzyme into an endo-β(1,4)-glucanase. This study demonstrates that experimental approaches can reveal structure-function relationships out of reach of current bioinformatic predictions.

KEYWORDS:

CAZyme; Saccharophagus degradans; crystallography; endoglucanase; enzyme; glycobiology; glycoside hydrolase; lichenase; oligomerization

PMID:
26755730
PMCID:
PMC4807298
DOI:
10.1074/jbc.M115.695999
[Indexed for MEDLINE]
Free PMC Article

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