PDBsum entry 3afb

Hydrolase

PDB id: 3afb

Contents

Protein chains

300 a.a. *

Ligands

GOL ×4

SO4

Metals

_MG ×2

Waters ×474

* Residue conservation analysis

PDB id:

3afb

Name:

Hydrolase

Title:

Crystal structures of catalytic site mutants of active domain 2 of chitinase from pyrococcus furiosus

Structure:

Putative chitinase. Chain: a, b. Fragment: catalytic domain (ad2), unp residues 409-717. Engineered: yes. Mutation: yes

Source:

Pyrococcus furiosus. Organism_taxid: 2261. Gene: pf1233. Expressed in: escherichia coli. Expression_system_taxid: 562

Resolution:

1.76Å R-factor: 0.162 R-free: 0.182

Authors:

H.Tsuji

Key ref:

H.Tsuji et al. (2010). Kinetic and crystallographic analyses of the catalytic domain of chitinase from Pyrococcus furiosus- the role of conserved residues in the active site. Febs J, 277, 2683-2695. PubMed id: 20553502

Date:

25-Feb-10 Release date: 09-Jun-10

Protein chains

Q8U1H5  (Q8U1H5_PYRFU) -  Chitinase from Pyrococcus furiosus (strain ATCC 43587 / DSM 3638 / JCM 8422 / Vc1)

Seq: 717 a.a.

Struc: 300 a.a.*

* PDB and UniProt seqs differ at 3 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.3.2.1.14 - chitinase.
• Reaction: Hydrolysis of the 1,4-beta-linkages of N-acetyl-D-glucosamine polymers of chitin.

Febs J 277:2683-2695 (2010)

PubMed id: 20553502

Kinetic and crystallographic analyses of the catalytic domain of chitinase from Pyrococcus furiosus- the role of conserved residues in the active site.

H.Tsuji, S.Nishimura, T.Inui, Y.Kado, K.Ishikawa, T.Nakamura, K.Uegaki.

ABSTRACT

The hyperthermostable chitinase from the hyperthermophilic archaeon Pyrococcus furiosus has a unique multidomain structure containing two chitin-binding domains and two catalytic domains, and exhibits strong crystalline chitin hydrolyzing activity at high temperature. In order to investigate the structure-function relationship of this chitinase, we analyzed one of the catalytic domains (AD2) using mutational and kinetic approaches, and determined the crystal structure of AD2 complexed with chito-oligosaccharide substrate. Kinetic studies showed that, among the acidic residues in the signature sequence of family 18 chitinases (DXDXE motif), the second Asp (D(2)) and Glu (E) residues play critical roles in the catalysis of archaeal chitinase. Crystallographic analyses showed that the side-chain of the catalytic proton-donating E residue is restrained into the favorable conformer for proton donation by a hydrogen bond interaction with the adjacent D(2) residue. The comparison of active site conformations of family 18 chitinases provides a new criterion for the subclassification of family 18 chitinase based on the conformational change of the D(2) residue.