Genomic characterization of methylotrophy of Oharaeibacter diazotrophicus strain SM30^T

Oharaeibacter diazotrophicus strain SM30^T, isolated from rice rhizosphere, is an aerobic, facultative lanthanide (Ln³⁺)-utilizing methylotroph and diazotroph that belongs to the Methylocystaceae family. In this research, the complete genome sequence of strain SM30^T was determined, and its methylotrophy modules were characterized. The genome consists of one chromosome and two plasmids, comprising a total of 5,004,097 bp, and the GC content was 71.6 mol%. A total of 4497 CDSs, 67 tRNA, and 9 rRNA were encoded. Typical alpha-proteobacterial methylotrophy genes were found: pyrroloquinoline quinone (PQQ)-dependent methanol dehydrogenase (MDH) (mxaF and xoxF1-4), methylotrophy regulatory proteins (mxbDM and mxcQE), PQQ synthesis, H₄F pathway, H₄MPT pathway, formate oxidation, serine cycle, and ethylmalonyl-CoA pathway. SDS-PAGE and subsequent LC–MS analysis, and qPCR analysis revealed that MxaF and XoxF1 were the dominant MDH in the absence or presence of lanthanum (La³⁺), respectively. The growth of MDH gene-deletion mutants on alcohols and qPCR results indicated that mxaF and xoxF1 are also involved in ethanol and propanol oxidation, xoxF2 participates in methanol oxidation in the presence of La³⁺, while xoxF3 was associated with methanol and ethanol oxidation in the absence of La³⁺, implying that XoxF3 is a calcium (Ca²⁺)-binding XoxF. Four Ln³⁺ such as La³⁺, cerium (Ce³⁺), praseodymium (Pr³⁺), and neodymium (Nd³⁺) served as cofactors for XoxF1 by supporting ΔmxaF growth on methanol. Some heavier lanthanides inhibited growth of SM30 on methanol. This study contributes to the understanding of the function of various XoxF-type MDHs and their roles in methylotrophs.