Co-localization of particulate methane monooxygenase and cd 1 nitrite reductase in the denitrifying methanotroph 'candidatus methylomirabilis oxyfera'

Candidatus Methylomirabilis oxyfera; is a polygon-shaped bacterium that was shown to have the unique ability to couple anaerobic methane oxidation to denitrification, through a newly discovered intra-aerobic pathway. Recently, the complete genome of Methylomirabilis oxyfera was assembled into a 2.7-Mb circular single chromosome by metagenomic sequencing. The genome of M.similar to oxyfera revealed the full potential to perform both methane oxidation and the conversion of nitrite via nitric oxide into oxygen and dinitrogen gas. In this study, we show by immunogold localization that key enzymes from both methane- and nitrite-converting pathways are indeed present in single M.similar to oxyfera cells. Antisera targeting the particulate methane monooxygenase (pMMO) and the cd1 nitrite reductase (NirS) were raised and used for immunogold localization in both single- and double-labelling experiments. Our previous studies have shown that M.similar to oxyfera does not develop pMMO-containing intracytoplasmic membranes as is observed in classical proteobacterial methanotrophs. Our results suggest that in M.similar to oxyfera, the pMMO and NirS enzymes localized to the cytoplasmic membrane and periplasm, respectively. Further, double-labelling showed co-occurrence of pMMO and NirS in single M.similar to oxyfera cells.