PDBsum entry 6kwc

Hydrolase

PDB id: 6kwc

Contents

Protein chain

191 a.a.

Ligands

GOL

Metals

IOD ×4

Waters ×260

References listed in PDB file

Key reference

• Title: Studying the role of a single mutation of a family 11 glycoside hydrolase using high-Resolution X-Ray crystallography.
• Authors: Z.Li, X.Zhang, C.Li, A.Kovalevsky, Q.Wan.
• Ref.: Protein J, 2020, 39, 671-680. [DOI no: 10.1016/j.polymdegradstab.2011.01.010]
• PubMed id: 33128114

Abstract

XynII is a family 11 glycoside hydrolase that uses the retaining mechanism for catalysis. In the active site, E177 works as the acid/base and E86 works as the nucleophile. Mutating an uncharged residue (N44) to an acidic residue (D) near E177 decreases the enzyme's optimal pH by ~ 1.0 unit. D44 was previously suggested to be a second proton carrier for catalysis. To test this hypothesis, we abolished the activity of E177 by mutating it to be Q, and mutated N44 to be D or E. These double mutants have dramatically decreased activities. Our high-resolution crystallographic structures and the microscopic pK_a calculations show that D44 has similar position and pK_a value during catalysis, indicating that D44 changes electrostatics around E177, which makes it prone to rotate as the acid/base in acidic conditions, thus decreases the pH optimum. Our results could be helpful to design enzymes with different pH optimum.