Bilirubin oxidase from Myrothecium verrucaria: X-ray determination of the complete crystal structure and a rational surface modification for enhanced electrocatalytic O2 reduction

被引:113
作者
Cracknell, James A. [1 ]
McNamara, Thomas P. [1 ]
Lowe, Edward D. [2 ]
Blanford, Christopher F. [1 ,3 ]
机构
[1] Univ Oxford, Inorgan Chem Lab, Dept Chem, Oxford OX1 3QR, England
[2] Univ Oxford, Dept Biochem, Oxford OX1 3QU, England
[3] Univ Manchester, Sch Mat, Manchester Interdisciplinary Bioctr, Manchester M13 9PL, Lancs, England
基金
英国工程与自然科学研究理事会;
关键词
DIRECT ELECTRON-TRANSFER; COPPER SITE; FUEL-CELLS; GRAPHITE; LACCASE; SUBSTRATE; T1; BIOELECTROCATALYSIS; DECOLORIZATION; PERTURBATIONS;
D O I
10.1039/c0dt01403f
中图分类号
O61 [无机化学];
学科分类号
070301 ; 081704 ;
摘要
The blue multi-copper oxidase bilirubin oxidase (BOx) from the ascomycete plant pathogen Myrothecium verrucaria (Mv) efficiently catalyses the oxidation of bilirubin to biliverdin, with the concomitant reduction of O-2 to water, a reaction of considerable interest for low-temperature bio-fuel cell applications. We have solved the complete X-ray determined structure of Mv BOx at 2.4 angstrom resolution, using molecular replacement with the Spore Coat Protein A (CotA) enzyme from Bacillus subtilis (PDB code 1GSK) as a template. The structure reveals an unusual environment around the blue type 1 copper (T1 Cu) that includes two non-coordinating hydrophilic amino acids, asparagine and threonine. The presence of a long, narrow and hydrophilic pocket near the T1 Cu suggests that structure of the substrate-binding site is dynamically determined in vivo. We show that the interaction between the binding pocket of Mv BOx and its highly conjugated natural organic substrate, bilirubin, can be used to stabilise the enzyme on a pyrolytic graphite electrode, more than doubling its electrocatalytic activity relative to the current obtained by simple adsorption of the protein to the carbon surface.
引用
收藏
页码:6668 / 6675
页数:8
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