Regulation of nitrogenase activity through iron protein interconversion into an active and an inactive form in Rhodopseudomonas capsulata

Abstract

The iron protein of nitrogenase from Rhodopseudomonas capsulata has been isolated in two forms depending on the nitrogen source used for growth. When isolated from cells grown on a limiting amount of ammonia, the Fe protein was composed of two equal subunits of apparent molecular weight 33 500. In contrast, the Fe protein purified from glutamate-grown cells was mainly inactive and composed of two different subunits of apparent M r 33 500 and 38 000. A two-dimensional immunoelectrophoresis technique was devised to allow direct determination, in cell-free extracts, of the subunit composition of Fe protein. By this method it is demonstrated that the Fe protein shifted from an active form (single type of subunit) to an inactive form (two distinct subunits) in cells submitted to an ammonia shock (15 mM NH 4Cl). Upon incubation of inactivated cells in a nitrogen-free medium, in vivo nitrogenase activity was recovered and, at the same time, the Fe protein was shifted back to the active form. This activation process was antagonized by chloramphenicol. In cells grown in the presence of [ 32P]phosphate, radioactivity was incorporated in the 38 kDa subunit of the inactive Fe protein. Upon treatment with the activating enzyme from Rhodospirillum rubrum, removal of radioactive phosphate was concomitant with the disappearance of the 38 kDa subunit. Altogether, these results suggest that nitrogenase activity in Rps. capsulata is controlled by the interconversion of the Fe protein between an active and an inactive form as in R. rubrum. This system of regulation appeared to turn off nitrogenase activity with more or less efficiency, depending on factors such as the nature and availability of the nitrogen source and the culture age.