Involvement of MmoR and MmoG in the transcriptional activation of soluble methane monooxygenase genes in Methylosinus trichosporium OB3b

Methanotrophs oxidize methane to methanol using the enzyme methane monooxygenase. Methylosinus trichosporium OB3b has two such enzymes: a membrane-bound particulate methane monooxygenase (pMMO) and a soluble, cytoplasmic methane monooxygenase (sMMO). In methanotrophs possessing both enzymes, the expression of the genes encoding sMMO and pMMO is regulated by copper ions, with sMMO expressed solely when copper is limiting. Virtually nothing is known about the specific machinery involved in the copper-regulated transcription of mmo genes except the identification of two proteins necessary for the expression: a sigma(54)-dependent transcriptional activator, MmoR, and a putative GroEL-like chaperone, MmoG. Genes encoding mmoR and mmoG are located immediately upstream of those encoding sMMO in the genome of M. trichosporium OB3b. Here, we use a green fluorescent protein promoter probe vector to show that nearly the complete intergenic DNA sequence between mmoG and mmoX is absolutely required for transcriptional activation. Furthermore, we used gel-shift assays to demonstrate that both MmoR and MmoG were required for protein binding to this region of DNA.