Draft genomes of gammaproteobacterial methanotrophs isolated from terrestrial ecosystems

Genome sequences of Methylobacter luteus, Methylobacter whittenburyi, Methylosarcina fibrata, Methylomicrobium agile, and Methylovulum miyakonense were generated. The strains represent aerobic methanotrophs typically isolated from various terrestrial ecosystems

Interchangeability of class I and II fumarases in an obligate methanotroph Methylotuvimicrobium alcaliphilum 20Z.

The methanotrophic bacterium Methylotuvimicrobium alcaliphilum 20Z is an industrially promising candidate for bioconversion of methane into value-added chemicals. Here, we have study the metabolic consequences of the breaking in the tricarboxylic acid (TCA) cycle by fumarase knockout. Two fumarases belonging to non-homologous class I and II fumarases were obtained from the bacterium by heterologous expression in Escherichia coli. Class I fumarase (FumI) is a homodimeric enzyme catalyzing the reversible hydration of fumarate and mesaconate with activities of ~94 and ~81 U mg-1 protein, respectively. The enzyme exhibited high activity under aerobic conditions, which is a non-typical property for class I fumarases characterized to date. The calculation of kcat/S0.5 showed that the enzyme works effectively with either fumarate or mesaconate, but it is almost four times less specific to malate. Class II fumarase (FumC) has a tetrameric structure and equal activities of both fumarate hydration and malate dehydration (~45 U mg-1 protein). Using mutational analysis, it was shown that both forms of the enzyme are functionally interchangeable. The triple mutant strain 20Z-3E (ΔfumIΔfumCΔmae) deficient in the genes encoding the both fumarases and the malic enzyme accumulated 2.6 and 1.1 mmol g-1 DCW fumarate in the medium when growing on methane and methanol, respectively. Our data suggest the redundancy of the metabolic node in the TCA cycle making methanotroph attractive targets for modification, including generation of strains producing the valuable metabolites

Methylotrophic Autotrophy in Beijerinckia mobilis

Representatives of the genus Beijerinckia are known as heterotrophic, dinitrogen-fixing bacteria which utilize a wide range of multicarbon compounds. Here we show that at least one of the currently known species of this genus, i.e., Beijerinckia mobilis, is also capable of methylotrophic metabolism coupled with the ribulose bisphosphate (RuBP) pathway of C(1) assimilation. A complete suite of dehydrogenases commonly involved in the sequential oxidation of methanol via formaldehyde and formate to CO(2) was detected in cell extracts of B. mobilis grown on CH(3)OH. Carbon dioxide produced by oxidation of methanol was further assimilated via the RuBP pathway as evidenced by reasonably high activities of phosphoribulokinase and ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO). Detection and partial sequence analysis of genes encoding the large subunits of methanol dehydrogenase (mxaF) and form I RubisCO (cbbL) provided genotypic evidence for methylotrophic autotrophy in B. mobilis