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Structural basis of chitin utilization by a GH20 β‐ N ‐acetylglucosaminidase from Vibrio campbellii strain ATCC BAA‐1116
Author(s) -
Meekrathok Piyanat,
Bürger Marco,
Porfetye Arthur T.,
Kumsaoad Sawitree,
Aunkham Anuwat,
Vetter Ingrid R.,
Suginta Wipa
Publication year - 2021
Publication title -
acta crystallographica section d
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.374
H-Index - 138
ISSN - 2059-7983
DOI - 10.1107/s2059798321002771
Subject(s) - chitin , mutant , active site , vibrio harveyi , stereochemistry , strain (injury) , chemistry , hydrolase , enzyme , biology , biochemistry , vibrio , bacteria , gene , genetics , anatomy , chitosan
Vibrio species play a crucial role in maintaining the carbon and nitrogen balance between the oceans and the land through their ability to employ chitin as a sole source of energy. This study describes the structural basis for the action of the GH20 β‐ N ‐acetylglucosaminidase ( Vh GlcNAcase) in chitin metabolism by Vibrio campbellii (formerly V. harveyi ) strain ATCC BAA‐1116. Crystal structures of wild‐type Vh GlcNAcase in the absence and presence of the sugar ligand, and of the unliganded D437A mutant, were determined. Vh GlcNAcase contains three distinct domains: an N‐terminal carbohydrate‐binding domain linked to a small α+β domain and a C‐terminal (β/α) 8 catalytic domain. The active site of Vh GlcNAcase has a narrow, shallow pocket that is suitable for accommodating a small chitooligosaccharide. Vh GlcNAcase is a monomeric enzyme of 74 kDa, but its crystal structures show two molecules of enzyme per asymmetric unit, in which Gln16 at the dimeric interface of the first molecule partially blocks the entrance to the active site of the neighboring molecule. The GlcNAc unit observed in subsite −1 makes exclusive hydrogen bonds to the conserved residues Arg274, Tyr530, Asp532 and Glu584, while Trp487, Trp546, Trp582 and Trp505 form a hydrophobic wall around the −1 GlcNAc. The catalytic mutants D437A/N and E438A/Q exhibited a drastic loss of GlcNAcase activity, confirming the catalytic role of the acidic pair (Asp437–Glu438).