Recovery of nitrogenase from aerobically isolated soybean nodule bacteroids

Bacteroids, the symbiotic forms of rhizobia, express nitrogenase, which is the enzyme that catalyzes the reduction of atmospheric dinitrogen to ammonium. The extreme oxygen lability of nitrogenase requires that bacteroids be isolated under anaerobic conditions to preserve their ability to reduce atmospheric dinitrogen. Aerobically isolated bacteroids were found to exhibit nitrogenase activity as measured via the acetylene reduction method but only after incubation of the bacteroids for about 1 h under low partial pressures of oxygen. The recovery of nitrogenase activity was dependent upon the partial pressure of oxygen to which the bacteroids were exposed during the one-hour incubation. The recovery of acetylene reduction activity was prevented by addition of inhibitors of protein synthesis, but not by inhibitors of transcription. Acetylene reduction could be expressed from aerobically isolated bacteroids stored for up to three days after isolation. The recovery of nitrogenase demonstrates that both the plant and the bacteroids possess mechanisms to protect the enzyme against oxygen. The recovery of nitrogen fixation activity from aerobically isolated bacteroids may provide a facile way to obtain bacteroids for studies on symbiotic functioning.