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Structure and function of Bs164 β-mannosidase from Bacteroides salyersiae the founding member of glycoside hydrolase family GH164

DOI: 10.1074/jbc.RA119.011591 DOI Help

Authors: Zachary Armstrong (University of York) , Gideon J. Davies (University of York)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Journal Of Biological Chemistry

State: Published (Approved)
Published: December 2019
Diamond Proposal Number(s): 18598

Open Access Open Access

Abstract: Recent work exploring protein sequence space has revealed a new glycoside hydrolase (GH) family (GH164) of putative mannosidases. GH164 genes are present in several commensal bacteria, implicating these genes in the degradation of dietary glycans. However, little is known about the structure, mechanism of action and substrate specificity of these enzymes. Herein we report the biochemical characterization and crystal structures of the founding member of this family (Bs164) from the human gut symbiont Bacteroides salyersiae. Previous reports of this enzyme indicated that it has α-mannosidase activity, however we conclusively show that it cleaves only β-mannose linkages. Using NMR spectroscopy, detailed enzyme kinetics of wild-type and mutant Bs164, and multi-angle light scattering we found that it is a trimeric retaining β-mannosidase, that is susceptible to several known mannosidase inhibitors. X-ray crystallography revealed the structure of Bs164 – the first known structure of a GH164 – at 1.91 Å resolution. Bs164 is composed of three domains: a (β/α)8 barrel, a trimerization domain and a β-sandwich domain, representing a previously unobserved structural fold for β-mannosidases. Structures of Bs164 at 1.80-2.55 Å resolution in complex with the inhibitors noeuromycin, mannoimidazole or DNP 2-deoxy-2-fluoro-mannose reveal the residues essential for specificity and catalysis including the catalytic nucleophile (Glu297) and acid/base residue (Glu160). These findings further our knowledge of the mechanisms commensal microbes use for nutrient acquisition.

Journal Keywords: Carbohydrate-active enzyme; Mannosidase; conformational analysis; Reaction Mechanism; structural biology; enzyme catalysis; glycoside hydrolase; glycosidase; carbohydrate chemistry; carbohydrate processing

Diamond Keywords: Enzymes; Bacteria; Gut Microbiota

Subject Areas: Biology and Bio-materials, Chemistry

Instruments: I04-Macromolecular Crystallography

Added On: 08/01/2020 12:19


Discipline Tags:

Catalysis Life Sciences & Biotech Structural biology Chemistry Biochemistry

Technical Tags:

Diffraction Macromolecular Crystallography (MX)