Rates of respiratory CO(2) loss and nitrogenase activities of H(2) uptake-negative mutant strains and H(2) uptake-positive revertant strains of Rhizobium japonicum have been investigated. Two-dimensional gel protein patterns of bacteroids formed by inoculation of soybeans (Glycine max L.) with these two strains show that they are closely related and revealed only one obvious difference between them. On the basis of molecular weight standards, it was concluded that the missing protein spot in the H(2) uptake-negative mutant strain could be caused by a failure of the mutant to synthesize hydrogenase. Nodules formed by the H(2) uptake-negative mutant strain evolved respiratory CO(2) at a rate of about 10% higher than that of nodules formed by the H(2) uptake-positive revertant strain. During short-term experiments employed, rates of both C(2)H(2) reduction and (15)N(2) fixation varied considerably among replicate samples and no statistically significant differences between mutant and revertant strains were observed. It was observed that increasing the partial pressure of O(2) over nodules significantly decreased the proportion of nitrogenase electrons allocated to H(+).