The mxaAKL genes of Methylobacter albus BG8

Abstract

The facultative methanol utilizer Methylobacterium extorquens AM1 contains at least three genes (mxaA, K and L) that encode functions involved in providing calcium to the holoenzyme of methanol dehydrogenase, the enzyme that oxidizes methanol to formaldehyde in this strain. Methane-utilizing bacteria (methanotrophs) also contain methanol dehydrogenase, and evidence suggests that similar methanol oxidation (Mox) functions may be present in some of these strains. DNA fragments from Methylobacterium extorquens AM1 specific to mxaA, mxaK and mxaL were isolated for use as hybridization probes against genomic digests of a variety of methanotrophic bacteria. Only the mxaL probe showed substantial hybridization, and it was used to identify and isolate an 8·5 kb HindIII fragment from Methylobacter albus BG8 (a Type I methanotroph). Hybridization of restriction digests of this fragment to individual probes for Methylobacterium extorquens AM1 mxaA, K and L indicated that the relative mxa gene order in Methylobacter albus BG8 is A-K-L. A T7 dual promoter/polymerase protein expression system indicated that five polypeptides are expressed from a 4·5 kb region of Methylobacter albus BG8 DNA in Escherichia coli, all transcribed in the same direction, and they apparently correspond to mxaACKDL. The functions of mxaC and mxaD are currently not known, but the order of mxaDL is reversed in Methylobacter albus BG8 compared to Methylobacterium extorquens AM1. When subclones o the Methylobacter albus BG8 fragment containing these genes were used as hybridization probes to genomic digests of methanotrophic bacteria, specific bands were detected that suggested a similar gene order in most cases. These data indicate that the mxaAKL region is relatively highly conserved in methanotrophs, and that in most cases the mxaAKL genes are grouped together in the same order as in the facultative methanol utilizer Methylobacterium extorquens AM1.