Glutamate biosynthesis in Bacillus azotofixans. 15N NMR and enzymatic studies.
Pathways of ammonia assimilation into glutamic acid in Bacillus azotofixans, a recently characterized nitrogen-fixing species of Bacillus, were investigated through observation by NMR spectroscopy of in vivo incorporation of 15N into glutamine and glutamic acid in the absence and presence of inhibitors of ammonia-assimilating enzymes, in combination with measurements of the specific activities of glutamate dehydrogenase, glutamine synthetase, glutamate synthase, and alanine dehydrogenase. In ammonia-grown cells, both the glutamine synthetase/glutamate synthase and the glutamate dehydrogenase pathways contribute to the assimilation of ammonia into glutamic acid. In nitrate-grown and nitrogen-fixing cells, the glutamine synthetase/glutamate synthase pathway was found to be predominant. NADPH-dependent glutamate dehydrogenase activity was detectable at low levels only in ammonia-grown and glutamate-grown cells. Thus, B. azotofixans differs from Bacillus polymyxa and Bacillus macerans, but resembles other N2-fixing prokaryotes studied previously, as to the pathway of ammonia assimilation during ammonia limitation. Implications of the results for an emerging pattern of ammonia assimilation by alternative pathways among nitrogen-fixing prokaryotes are discussed, as well as the utility of 15N NMR for measuring in vivo glutamate synthase activity in the cell.
D000641:Ammonia / Q000378:metabolism; D001407:Bacillus / Q000378:metabolism*; D005970:Glutamate Synthase / Q000378:metabolism; D005971:Glutamates / Q000096:biosynthesis*; D018698:Glutamic Acid; D009682:Magnetic Resonance Spectroscopy; D009243:NAD / Q000378:metabolism; D009249:NADP / Q000378:metabolism; D009586:Nitrogen Fixation; D013997:Time Factors
D005971:Glutamates; D009243:NAD; D018698:Glutamic Acid; D009249:NADP; D000641:Ammonia; D005970:Glutamate Synthase