Mutants of Azotobacter which grow normally on excess ammonia under a variety of conditions and which grow slowly or not at all on atmospheric nitrogen have been isolated. Extracts of these strains have low or no detectable nitrogenase activity. There are three classes of mutants. Cell-free preparations of members of the first class possess an enhancement factor (EF+) which stimulates wild-type nitrogenase
in vitro. Homogenates of members of the second class possess an enhanceable factor (EF-) which complements in vitrowith extracts of the first class of mutants to give substantial nitrogenase activity. Preparations of members of the third class contain neither EF+ nor EF- activity. EF+ and EF- are repressed by the same conditions that repress nitrogenase. Molybdenum-deficient cells of the second class of mutants do not appear to contain EF- activity, but molybdenum deficient cells of the first class of mutants contain EF+. Because of these observations, EF+ is tentatively equated to azoferredoxin and EF- to molybdoferredoxin.