Glucose oxidase

 

Glucose oxidase is a flavin dependent glycoprotein. The fungal enzyme is a homodimer made up of two identical subunits, each containing one molecule of non-covalently bound FAD. The enzyme catalyses the oxidation of beta-D-glucose, where the FAD cofactor acts as the redox carrier.

 

Reference Protein and Structure

Sequence
P13006 UniProt (1.1.3.4) IPR012132 (Sequence Homologues) (PDB Homologues)
Biological species
Aspergillus niger (Fungus) Uniprot
PDB
1gal - CRYSTAL STRUCTURE OF GLUCOSE OXIDASE FROM ASPERGILLUS NIGER: REFINED AT 2.3 ANGSTROMS RESOLUTION (2.3 Å) PDBe PDBsum 1gal
Catalytic CATH Domains
3.30.560.10 CATHdb (see all for 1gal)
Cofactors
Fadh2(2-) (1)
Click To Show Structure

Enzyme Reaction (EC:1.1.3.4)

beta-D-glucose
CHEBI:15903ChEBI
+
dioxygen
CHEBI:15379ChEBI
→
hydrogen peroxide
CHEBI:16240ChEBI
+
D-glucono-1,5-lactone
CHEBI:16217ChEBI
Alternative enzyme names: Beta-D-glucose oxidase, Beta-D-glucose:quinone oxidoreductase, D-glucose oxidase, D-glucose-1-oxidase, GOD, Corylophyline, Deoxin-1, Glucose aerodehydrogenase, Glucose oxyhydrase, Microcid, Penatin, Beta-D-glucose:oxygen 1-oxido-reductase,

Enzyme Mechanism

Introduction

Crystallographic investigations have shown the flavin containing active site to be buried in a deep pocket. The binding of glucose to the free enzyme results in the expulsion of a water molecule and a proton from the active site.

In the reduction half reaction, simultaneous hydride and proton transfer from the glucose to the FAD and His516 respectively occurs.

In the oxidation half reaction, the reduced coenzyme FADH- is reoxidised back to FAD by molecular dioxygen which undergoes reduction to hydrogen peroxide in two single electron transfer steps.

The hydride transfer is thought to be regulated by dissociation between Glu412 and His559. The electrostatic interaction between these residues controls the pH and therefore reactivity of the active site, although they are not directly involved in the catalytic mechanism.

Catalytic Residues Roles

UniProt PDB* (1gal)
His538 His516A The residue acts as a general base towards the C1-OH group of the glucose substrate. As the proton is removed, the oxygen partial negative charge drives the hydride transfer to the N5 of FAD resulting in FADH-. A ketone is formed at the C1 of glucose. proton acceptor
*PDB label guide - RESx(y)B(C) - RES: Residue Name; x: Residue ID in PDB file; y: Residue ID in PDB sequence if different from PDB file; B: PDB Chain; C: Biological Assembly Chain if different from PDB. If label is "Not Found" it means this residue is not found in the reference PDB.

Chemical Components

hydride transfer, proton transfer, native state of cofactor regenerated, overall reactant used, aromatic bimolecular nucleophilic addition, overall product formed

References

  1. Leskovac V et al. (2005), Int J Biochem Cell Biol, 37, 731-750. Glucose oxidase from Aspergillus niger: the mechanism of action with molecular oxygen, quinones, and one-electron acceptors. DOI:10.1016/j.biocel.2004.10.014. PMID:15694834.
  2. Wang Y et al. (2018), Chem Sci, 9, 594-599. FAD roles in glucose catalytic oxidation studied by multiphase flow of extractive electrospray ionization (MF-EESI) mass spectrometry. DOI:10.1039/C7SC04259K. PMID:29629123.
  3. Hecht HJ et al. (1993), J Mol Biol, 229, 153-172. Crystal structure of glucose oxidase from Aspergillus niger refined at 2.3 A resolution. PMID:8421298.

Step 1. In the reductive half-reaction there is concerted transfer of a proton from glucose to His516 and a hydride from glucose to FAD, forming FADH- and the product. FADH- is reoxidized by oxygen in the oxidative half-reaction. This proceeds via two single-electron transfer steps and involves superoxide anion radical and flavin semiquinone radical intermediates.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
His516A proton acceptor

Chemical Components

hydride transfer, proton transfer, native state of cofactor regenerated, overall reactant used, ingold: aromatic bimolecular nucleophilic addition, overall product formed
Download: Image, Marvin File

Step 1. In the reductive half-reaction there is concerted transfer of a proton from glucose to His516 and a hydride from glucose to FAD, forming FADH- and the product. FADH- is reoxidized by oxygen in the oxidative half-reaction. This proceeds via two single-electron transfer steps and involves superoxide anion radical and flavin semiquinone radical intermediates.

Products.

Contributors

James W. Murray, Craig Porter, Gemma L. Holliday, Amelia Brasnett