N-2 FIXATION IN MARINE HETEROTROPHIC BACTERIA - DYNAMICS OF ENVIRONMENTAL AND MOLECULAR REGULATION

Molecular and immunological techniques were used to examine N-2 fixati on in a ubiquitous heterotrophic marine bacterium, the facultative ana erobic Vibrio natriegens. When batch cultures were shifted from aerobi c N-replete to anaerobic N-deplete conditions, transcriptional and pos ttranslational regulation of N-2 fixation was observed, Levels of nifH DK mRNA encoding the nitrogenase enzyme mere highest at 140 min postsh ift and undetectable between 6 and 9 h later, Immunologically determin ed levels of nitrogenase enzyme (Fe protein) were highest between 6 an d 15 h postshift, and nitrogenase activity peaked between 6 and 9 h po stshift, declining by a factor of 2 after 12-15 h, Unlike their regula tion in cyanobacteria, Fe protein and nitrogenase activity were presen t when nifHDK mRNA was absent in V. natriegens, indicating that nitrog enase is stored and stable under anaerobic conditions, Both nifHDK mRN A and Fe protein disappeared within 40 min after cultures were shifted from N2-fixing conditions (anaerobic, N-deplete) to non-N-2-fixing co nditions (aerobic, N-enriched) but reappeared when shifted to conditio ns favoring N-2 fixation, Thus, unlike other N-2-fixing heterotrophic bacteria, nitrogenase must be resynthesized after aerobic exposure in V. natriegens. Immunological detection based on immunoblot (Western) a nalysis and immunogold labeling correlated positively with nitrogenase activity; no localization of nitrogenase was observed, Because V. nat riegens continues to fix N-2 for many hours after anaerobic induction, this species may play an important role in providing ''new'' nitrogen in marine ecosystems.