Effect of Ammonia on the Synthesis and Function of the N2-Fixing Enzyme System in Clostridium pasteurianum

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The N2-fixing system of Clostridium pasteurianum operates under regulatory controls; no activity is found in cultures growing on excess NH3. The conditions which are necessary for the synthesis and function of this system were studied in whole cells by using acetylene reduction as a sensitive assay for the presence of the N2-fixing system. Nitrogenase of N2-fixing cultures normally can fix twice as much N2 as is needed to maintain the growth rate. When cultures that have grown for four or more generations on NH3 exhaust NH3 from the medium, a diauxic lag of about 90 min ensues before growth is resumed on N2. Neither N2-fixing nor acetylene reduction activity can be detected before growth is resumed on N2. N2 is not a necessary requirement for this synthesis since under argon that contains less than 10−8m N2, the N2-fixing system is made. If NH3 is added to N2-dependent cultures, synthesis of the enzyme system is abruptly stopped, but the enzyme already present remains stable and functional for at least 6 hr (over three generations). Cultures grown under argon in a chemostat controlled by limiting ammonia have derepressed nitrogenase synthesis. If the argon is removed and replaced by N2, partial repression of nitrogenase occurs.

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