EFFECTS OF AMMONIA ON THE DE-NOVO SYNTHESIS OF POLYPEPTIDES IN CELLS OF NITROSOMONAS-EUROPAEA DENIED AMMONIA AS AN ENERGY-SOURCE

The effects of ammonium on the de novo synthesis of polypeptides in th e soil-nitrifying bacterium Nitrosomonas europaea have been investigat ed. Cells were incubated in the presence of both acetylene and NH4+. U nder these conditions, the cells were unable to utilize NH4+ as an ene rgy source. Energy to support protein synthesis was supplied by the ox idation of hydroxylamine or other alternative substrates for hydroxyla mine oxidoreductase. De novo protein synthesis was detected by C-14 in corporation from (CO2)-C-14 into polypeptides by sodium dodecyl sulfat e-polyacrylamide gel electrophoresis and fluorography. In the presence of NH4+, acetylene-treated cells synthesized the 27-kDa polypeptide o f ammonia monooxygenase (AMO) and two other major polypeptides (with s izes of 55 and 65 kDa). The synthesis of these polypeptides was comple tely inhibited by chloramphenicol and attenuated by rifampin. The opti mal concentration of hydroxylamine for the in vivo C-14-labeling react ion was found to be 2 mM. The effect of NH4+ concentration was also ex amined. It was shown to cause a saturable response with a K-s of appro ximately 2.0 mM NH4+. Labeling studies conducted at different pH value s suggest cells respond to NH3 rather than NH4+. No other compounds te sted were able to influence the synthesis of the 27-kDa component of A MO, although we have also demonstrated that this polypeptide can be sy nthesized under anaerobic conditions in cells utilizing pyruvate- or h ydrazine-dependent nitrite reduction as an energy source. We conclude that ammonia has a regulatory effect on the synthesis of a subunit of AMO in addition to providing nitrogen for protein synthesis.