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New insights into the oxidative reaction of oxidases using copper amine oxidases as a model

Reference	BB/C00468X/1
Principal Investigator / Supervisor	Professor Michael McPherson
Co-Investigators / Co-Supervisors	Professor Simon Phillips
Institution	University of Leeds
Department	Inst of Molecular & Cellular Biology
Funding type	Research
Value (£)	232,339
Status	Completed
Type	Research Grant
Start date	01/09/2005
End date	30/09/2008
Duration	37 months

Abstract

We believe that phenomena related to the interaction of diatomic substrates and products and proton transfer steps in amine oxidases provide a simpler model for understanding such processes in more complex enzyme systems such as cytochrome c oxidase. Copper amine oxidases catalyse two distinct reactions. First, the single turnover generation of a trihydroxyphenylalanine (TPQ) cofactor by the insertion of oxygens into an active site tyrosine residue and second, the multiple turnover of amine substrate to aldehyde product with associated reoxidation of the enzyme releasing ammonium and hydrogen peroxide. There is good evidence that the formation of the cofactor involves redox activation of oxygen for insertion to form the TPQ moiety. Key questions are how oxygen enters and interacts at the active site of the enzyme during catalytic turnover, how product peroxide is released in a controlled manner and whether the copper plays a redox role during turnover. There is controversy based on previous reports of metal substitution in amine oxidases, in one case indicating copper is essential and in another that substitution by cobalt supports activity. We have recently demonstrated that recovery of activity in metal substituted E. coli enzyme is dependent on peripheral calciums, whose function has until now been unclear. These results support the view that copper plays a stabilising and not a redox role during turnover. We have developed a hypothesis that oxygen enters via a hydrophobic tunnel regulated by the calciums. We wish to test this hypothesis and to undertake comparative studies with other amine oxidases to investigate the role of the copper during the oxidative half reaction.

Summary

unavailable

Committee	Closed Committee - Biomolecular Sciences (BMS)
Research Topics	Industrial Biotechnology, Microbiology, Structural Biology
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

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