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Title: Regulation of methane monooxygenase genes in methanotrophs
Author: Stafford, Graham P.
ISNI:       0000 0001 3476 7697
Awarding Body: University of Warwick
Current Institution: University of Warwick
Date of Award: 2002
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Methanotrophs are bacteria capable of using methane as their sole carbon and energy source. Oxidation of methane by Methylosinus trichosporium OB3b is catalysed by the particulate (pMMO) and soluble (sMMO) methane monooxygenase enzymes. Transcription of the genes encoding pMMO (pmo) occurs at high copper-to-biomass ratio and sMMO (mmo) at low copper-to-biomass ratios. The aim of this work was to study the regulation of methane monooxygenase gene expression in response to copper ions- the "copper switch". Primer extension and RT-PCR revealed putative transcriptional start sites 5' of the pmoC genes from Methylocystis sp. strain M and Methylosinus trichosporium OB3b that corresponded to a"-promoters. The pmo operons from both organisms possessed tandem promoters 5' of pmoC suggesting differential expression of the two sets of pmo genes. A qualitative analysis of pmo transcription from M trichosporium OB3b revealed that the pmo operon is transcribed at both high and low copper ion concentrations when M. trichosporium OB3b was grown on methane or methanol. The gene encoding a" (rpoN) was cloned and sequenced, since previously Nielsen et al., (Mo! Microbiol (1997) 25, 399-409) had shown that transcription of the mmo genes from M trichosporium OB3b was shown to proceed from a a"-type promoter. Marker-exchange mutagenesis of rpoN verified its role in transcription of the mmo operon and also suggested its requirement for transcription of both nitrogen fixation and nitrate reduction genes. Sequencing of the region 5' of the mmo cluster (Ian McDonald, unpublished) revealed a gene possessing homology with a"-dependent transcriptional activators, named mmoR. Mutation of mmoR by marker-exchange mutagenesis showed it was also required for expression of the mmo operon. These data have allowed an improved model for the regulation of the pmo and mmo gene clusters by copper to be proposed.
Supervisor: Not available Sponsor: Biotechnology and Biological Sciences Research Council ; Society for General Microbiology ; European Research Council
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: QR Microbiology