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. 1978 Jul;75(7):3258–3262. doi: 10.1073/pnas.75.7.3258

Expression of hydrogenase activity in free-living Rhizobium japonicum

Robert J Maier *, Norman E R Campbell †, Frank J Hanus *, Frank B Simpson *, Sterling A Russell *, Harold J Evans *
PMCID: PMC392754  PMID: 16592544

Abstract

A medium is described on which selected Rhizobium japonicum strains express hydrogenase (H2 uptake) activity under free-living conditions. Low concentrations of carbon substrates, decreased oxygen tension, and the quantity of combined nitrogen in the medium were major factors influencing hydrogenase expression. Hydrogenase activity was dependent upon a preincubation period in the presence of H2 under conditions such that the cells did not exhibit nitrogenase activity. H2 uptake rates were easily measured amperometrically in aerobically or anaerobically prepared suspensions from free-living cultures. Six R. japonicum strains that formed nodules with the ability to utilize H2 oxidized this gas when grown in free-living cultures. In comparison six randomly chosen strains forming nodules that lost H2 in air either showed no or low capacity to take up H2 under free-living conditions. The reduction of triphenyltetrazolium chloride in an agar medium was used to detect strains capable of oxidizing H2. This method has enabled us to isolate a spontaneous R. japonicum mutant strain that has lost the ability to utilize H2. This mutant strain forms nodules that evolve H2 but other symbiotic characteristics appear normal. This strain will be useful in evaluating the importance of the hydrogenase system in the nitrogen-fixing process of legumes.

Keywords: nitrogen fixation, legumes, soybeans, mutant

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

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