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GmcA is a FAD-containing enzyme belonging to the GMC (glucose-methanol-choline oxidase) family of oxidoreductases. A mutation in the  Rhizobium leguminosarum gmcA  gene was generated by homologous recombination. The mutation in  gmcA  did not affect the growth of  R. leguminosarum , but it displayed decreased antioxidative capacity at H 2 O 2  conditions higher than 5 mM. The  gmcA  mutant strain displayed no difference of glutathione reductase activity, but significantly lower level of the glutathione peroxidase activity than the wild type. Although the  gmcA  mutant was able to induce the formation of nodules, the symbiotic ability was severely impaired, which led to an abnormal nodulation phenotype coupled to a 30% reduction in the nitrogen fixation capacity. The observation on ultrastructure of 4-week pea nodules showed that the mutant bacteroids tended to start senescence earlier and accumulate poly-β-hydroxybutyrate (PHB) granules. In addition, the  gmcA  mutant was severely impaired in rhizosphere colonization. Real-time quantitative PCR showed that the  gmcA  gene expression was significantly up-regulated in all the detected stages of nodule development, and statistically significant decreases in the expression of the redoxin genes  katG, katE , and  ohrB  were found in  gmcA  mutant bacteroids. LC-MS/MS analysis quantitative proteomics techniques were employed to compare differential  gmcA  mutant root bacteroids in response to the wild type infection. Sixty differentially expressed proteins were identified including 33 up-regulated and 27 down-regulated proteins. By sorting the identified proteins according to metabolic function, 15 proteins were transporter protein, 12 proteins were related to stress response and virulence, and 9 proteins were related to transcription factor activity. Moreover, nine proteins related to amino acid metabolism were over-expressed.

A GMC Oxidoreductase GmcA Is Required for Symbiotic Nitrogen Fixation in Rhizobium leguminosarum bv. viciae