PDBsum entry 2eie

Oxidoreductase

PDB id: 2eie

Contents

Protein chain

639 a.a. *

Ligands

AZI

Metals

_CU

Waters ×545

* Residue conservation analysis

PDB id:

2eie

Name:

Oxidoreductase

Title:

Crystal structure of galactose oxidase complexed with azide

Structure:

Galactose oxidase. Chain: a. Fragment: residues 1-639. Synonym: gao. Engineered: yes

Source:

Gibberella zeae. Organism_taxid: 5518. Expressed in: emericella nidulans. Expression_system_taxid: 162425.

Resolution:

1.80Å R-factor: 0.192 R-free: 0.209

Authors:

S.E.Phillips,M.J.Mcpherson,P.F.Knowles,N.Akyumani,S.J.Firbank, S.Tamber

Key ref:

M.S.Rogers et al. (2007). The stacking tryptophan of galactose oxidase: a second-coordination sphere residue that has profound effects on tyrosyl radical behavior and enzyme catalysis. Biochemistry, 46, 4606-4618. PubMed id: 17385891

Date:

12-Mar-07 Release date: 24-Apr-07

Protein chain

P0CS93  (GAOA_GIBZA) -  Galactose oxidase from Gibberella zeae

Seq: 680 a.a.

Struc: 639 a.a.

Enzyme reactions

• Enzyme class: E.C.1.1.3.9 - galactose oxidase.
• Reaction: D-galactose + O2 = D-galacto-hexodialdose + H2O2
• Cofactor: Cu cation

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

reference

Biochemistry 46:4606-4618 (2007)

PubMed id: 17385891

The stacking tryptophan of galactose oxidase: a second-coordination sphere residue that has profound effects on tyrosyl radical behavior and enzyme catalysis.

M.S.Rogers, E.M.Tyler, N.Akyumani, C.R.Kurtis, R.K.Spooner, S.E.Deacon, S.Tamber, S.J.Firbank, K.Mahmoud, P.F.Knowles, S.E.Phillips, M.J.McPherson, D.M.Dooley.

ABSTRACT

The function of the stacking tryptophan, W290, a second-coordination sphere residue in galactose oxidase, has been investigated via steady-state kinetics measurements, absorption, CD and EPR spectroscopy, and X-ray crystallography of the W290F, W290G, and W290H variants. Enzymatic turnover is significantly slower in the W290 variants. The Km for D-galactose for W290H is similar to that of the wild type, whereas the Km is greatly elevated in W290G and W290F, suggesting a role for W290 in substrate binding and/or positioning via the NH group of the indole ring. Hydrogen bonding between W290 and azide in the wild type-azide crystal structure are consistent with this function. W290 modulates the properties and reactivity of the redox-active tyrosine radical; the Y272 tyrosyl radicals in both the W290G and W290H variants have elevated redox potentials and are highly unstable compared to the radical in W290F, which has properties similar to those of the wild-type tyrosyl radical. W290 restricts the accessibility of the Y272 radical site to solvent. Crystal structures show that Y272 is significantly more solvent exposed in the W290G variant but that W290F limits solvent access comparable to the wild-type indole side chain. Spectroscopic studies indicate that the Cu(II) ground states in the semireduced W290 variants are very similar to that of the wild-type protein. In addition, the electronic structures of W290X-azide complexes are also closely similar to the wild-type electronic structure. Azide binding and azide-mediated proton uptake by Y495 are perturbed in the variants, indicating that tryptophan also modulates the function of the catalytic base (Y495) in the wild-type enzyme. Thus, W290 plays multiple critical roles in enzyme catalysis, affecting substrate binding, the tyrosyl radical redox potential and stability, and the axial tyrosine function.