2 DIFFERENTIALLY REGULATED NITRATE REDUCTASES REQUIRED FOR NITRATE-DEPENDENT, MICROAEROBIC GROWTH OF BRADYRHIZOBIUM-JAPONICUM

Native PAGE of Triton X-100-solubilized membranes from Bradyrhizobium japonicum strain PJ17 grown microaerobically (2% O-2, v/v) in defined nitrate-containing medium resolved two catalytically active nitrate re ductase (NR) species with apparent molecular masses of 160kDa (NR(I)) and 200kDa (NR(II)). NR(I) and NR(II) were also found in membranes fro m cells of strain PJ17 that were first grown in defined medium with gl utamate and further incubated microaerobically in the presence of 5 mm ol/l KNO3. However, only NR(I) was detected in cell membranes of strai n PJ17 when nitrate was omitted from the microaerobic incubation mediu m. Four mutants unable to grow at low O-2 tension in the presence of n itrate were isolated after transposon Tn5 muta,genesis. Membranes from mutants GRF110 and GRF116 showed mainly NR(I), while the other two mu tants, GRF3 and GRF4, expressed mostly NR(II). These results indicate that the ability of B. japonicum PJ17 to grow under microaerobic condi tions depends upon the presence of two membrane-bound NR enzymes whose synthesis seem to be independently induced by microaerobiosis (NR(I)) or by both microaerobiosis and nitrate (NR(II)).