Award details

Exploring the catalytic versatility of soluble methane monooxygenase

Reference	BB/C003276/1
Principal Investigator / Supervisor	Professor John Murrell
Co-Investigators / Co-Supervisors	Professor Thomas Smith
Institution	University of Warwick
Department	Biological Sciences
Funding type	Research
Value (£)	189,661
Status	Completed
Type	Research Grant
Start date	01/08/2004
End date	31/07/2007
Duration	36 months

Abstract

Soluble methane monooxygenase (sMMO) is a non-haem binuclear iron active-centre enzyme complex that oxygenates methane and numerous adventitious substrates. This proposal will investigate the structural determinants of methane-oxidising activity and the breadth of the sMMO substrate profile, as well as developing mutant enzymes and a new expression system for bioremediation of polyaromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs) and trichloroethene (TCE). sMMO is naturally produced by methanotrophic bacteria such as Methylosinus trichosporium, in which it catalyses the conversion of the unreactive hydrocarbon methane to methanol. Site-directed and saturation mutagenesis of key amino acyl residues, identified by comparison to the sequence of a homologous enzyme that does not oxidise methane, and will be used to identify the structural determinant(s) of methane oxidation. The substrate range of sMMO is much broader than those of its homologues and includes alkanes, alkenes, mono- and di-aromatics and the important groundwater pollutant trichloroethene. Saturation mutagenesis of one established crucial site, together with random mutagenesis of the active centre and surrounding regions, will be used to generate libraries of mutants for screening with a panel of key substrates. This will identify the amino acyl residues important in determining the uniquely broad substrate spectrum of sMMOs and generate novel enzymes with enhanced activities towards some of the most problematic environmental contaminants. The original methanotrophic bacterium, which is widespread in the environment, constitutes an excellent host for expression of recombinant enzymes for bioremediation applications and we already have a facile expression system for this organism that we have exploited in the laboratory. In order to make this system suitable to applications in the field, we will develop new, constitutive promoter system to produce recombinant sMMO expression that is relatively insensitive to environmental conditions and will maximise the impact of our novel enzymes in future bioremediation applications. (Joint with BB/C00339X/1).

Summary

unavailable

Committee	Closed Committee - Engineering & Biological Systems (EBS)
Research Topics	X – not assigned to a current Research Topic
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

Associated awards:

BB/C00339X/1 Exploring the catalytic versatility of soluble methane monooxygenase

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