Abstract
Ubiquinone (UQ) has been considered as an electron mediator in electron transfer that generates ATP in Rhizobium under both free-living and symbiosis conditions. When mutated, the dmtH gene has a symbiotic phenotype of forming ineffective nodules on Astragalus sinicus. The gene was isolated from a Mesorhizobium huakuii 7653R transposon-inserted mutant library. The DNA sequence and conserved protein domain analyses revealed that dmtH encodes demethylmenaquinone (DMK) methyltransferase, which catalyzes the terminal step of menaquinone (MK) biosynthesis. Comparative analysis indicated that dmtH homologs were present in only a few Rhizobia. Real-time quantitative PCR showed dmtH is a bacteroid-specific gene. The highest expression was seen at 25 days after inoculation of strain 7653R. Gene disruption and complementation tests demonstrated that the dmtH gene was essential for bacteroid development and symbiotic nitrogen fixation ability. MK and UQ were extracted from the wild type strain 7653R and mutant strain HK116. MK-7 was accumulated under microaerobic condition and UQ-10 was accumulated under aerobic condition in M. huakuii 7653R. The predicted function of DmtH protein was confirmed by the measurement of methyltransferase activity in vitro. These results revealed that MK-7 was used as an electron carrier instead of UQ in M. huakuii 7653R bacteroids.
Highlights
In bioenergetic chains, diverse chemical types of quinones, such as ubi, plasto, mena, rhodo, caldariella- or sulfolobus-quinones have been identified as membrane-bound, mobile hydrogen carriers in different species
It was deduced that the evolutionary transition of bioenergetic chains from high-potential MK to low-potential UQ in the proteobacterial phylum had drived by rising levels of dioxygen 2.5 billion years ago, and bioenergetic ambivalence of the respective organisms which work both on MK-and on UQ-pools, was necessary in transition proceed [5]
The homology analysis of the DmtH protein showed that had 99% identity to the function-unknown gene mll2332 from M. loti MAFF303099, and another similar proteins were annotated as demethylmenaquinone methyltransferase proteins in M. opportunistum WSM2075 (94% identity), R. leguminosarum bv. trifolii WSM1325 (90% identity) and R. etli CFN42 (88% identity)
Summary
Diverse chemical types of quinones, such as ubi-, plasto-, mena-, rhodo-, caldariella- or sulfolobus-quinones have been identified as membrane-bound, mobile hydrogen carriers in different species. Ubiquinone (UQ) is the dominant quinone species in a/b/c-proteobacteria. The majority of strictly aerobic Gram-negative bacteria contain exclusively UQ [1,2,3]. Menaquinone (MK) is most widely distributed in most Grampositive bacteria and anaerobic Gram-negative bacteria. E. coli and closely related species can switch between MK-and UQ-based bioenergetic chains in response to varying growth conditions. UQ-8 is used as the main quinone in aerobic respiration [6]. Under anaerobic conditions, MK-8 and demethylmenaquinone-8 (DMK-8) are synthesized as electron transferring quinones [7,8]
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