PDBsum entry 3a3v

Hydrolase

PDB id: 3a3v

Contents

Protein chain

376 a.a. *

Ligands

GOL ×4

Metals

_NI

Waters ×516

* Residue conservation analysis

PDB id:

3a3v

Name:

Hydrolase

Title:

Crystal structure of reducing-end-xylose releasing exo-oligoxylanase y198f mutant

Structure:

Xylanase y. Chain: a. Synonym: reducing-end-xylose releasing exo-oligoxylanase. Engineered: yes. Mutation: yes

Source:

Bacillus halodurans. Organism_taxid: 86665. Strain: c-125. Gene: bh2105. Expressed in: escherichia coli bl21. Expression_system_taxid: 511693.

Resolution:

1.39Å R-factor: 0.178 R-free: 0.198

Authors:

M.Hidaka,S.Fushinobu,Y.Honda,M.Kitaoka

Key ref:

M.Hidaka et al. (2010). Structural explanation for the acquisition of glycosynthase activity. J Biochem (tokyo), 147, 237-244. PubMed id: 19819900

Date:

22-Jun-09 Release date: 03-Nov-09

Protein chain

Q9KB30  (REOX_BACHD) -  Reducing end xylose-releasing exo-oligoxylanase from Halalkalibacterium halodurans (strain ATCC BAA-125 / DSM 18197 / FERM 7344 / JCM 9153 / C-125)

Seq: 388 a.a.

Struc: 376 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

Enzyme reactions

• Enzyme class: E.C.3.2.1.156 - oligosaccharide reducing-end xylanase.

J Biochem (tokyo) 147:237-244 (2010)

PubMed id: 19819900

Structural explanation for the acquisition of glycosynthase activity.

M.Hidaka, S.Fushinobu, Y.Honda, T.Wakagi, H.Shoun, M.Kitaoka.

ABSTRACT

Glycosynthases are engineered glycoside hydrolases (GHs) that catalyse the synthesis of glycoside from glycosyl-fluoride donors and suitable acceptors. We have determined five crystal structures of the glycosynthase mutants reducing-end xylose-releasing exo-oligoxylanase, an inverting GH, that exhibit various levels of glycosynthetic activities. At the active site of the Y198F mutant, the most efficient glycosynthase, a water molecule is observed at the same position as nucleophilic water (NW) in the parent enzyme, and the loss of the fixation of the direction of the lone pair of water molecules in the mutant drastically decreases hydrolytic activity. Water molecules were also observed at each active site of the general base mutant, but they were shifted 1.0-3.0 A from the NW in the wild type. Their positions exhibited a strong correlation with the strength of glycosynthase activity. Here, we propose that a structural prerequisite for the sufficient glycosynthase reaction is the presence of a water molecule at the NW position, and mutation at the NW holder provides a general strategy for inverting GHs. The idea on the position of a water molecule may also be applicable to the design of efficient glycosynthases from retaining GHs.