NifL of Klebsiella pneumoniae: redox characterization in relation to the nitrogen source.
In Klebsiella pneumoniae, NifL modulates the activity of the transcriptional activator NifA in response to combined nitrogen or external molecular oxygen. We recently showed that K. pneumoniae NifL is a flavoprotein which apparently senses oxygen through a redox-sensitive, conformational change. In order to study whether the nitrogen signal might be transmitted to NifA through a stable modification of NifL we characterized the redox properties of NifL synthesized in Escherichia coli in the presence of different nitrogen sources. FAD analyses showed that purified NifL carried FAD as cofactor independent of nitrogen and oxygen availability. The redox potential of NifL synthesized in the presence of ammonium was -277+/-5 mV at pH 8.0 and 25 degrees C, as determined by reduction with dithionite or with enzymatic reduction by xanthine oxidase in the presence of methyl viologen as redox mediator. When synthesized under nitrogen-limiting conditions, NifL showed a redox potential of -274+/-6 mV at pH 8.0 and 25 degrees C. Fully reduced NifL fractions, synthesized under either condition listed above, reoxidized rapidly in the presence of molecular oxygen. These results indicate that for NifL synthesized in E. coli, the redox potential of the NifL-bound FAD is not influenced by the nitrogen source. The two NifL fractions differed, however, in that a non-flavin specific absorbance at 420 nm was found only in NifL synthesized in the presence of ammonium.
Klopprogge
K
K
Institut für Mikrobiologie und Genetik, Universität Göttingen, Grisebachstr. 8, 37077, Göttingen, Germany.
Schmitz
R A
RA
D000641:Ammonia; D001059:Apoproteins / Q000737:chemistry; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D005182:Flavin-Adenine Dinucleotide / Q000737:chemistry; D002479:Inclusion Bodies / Q000378:metabolism; D007501:Iron / Q000032:analysis; D007711:Klebsiella pneumoniae / Q000378:metabolism*; D009586:Nitrogen Fixation; D010084:Oxidation-Reduction; D010544:Peroxidases / Q000032:analysis; D010957:Plasmids; D015398:Signal Transduction
D001059:Apoproteins; D001426:Bacterial Proteins; C084296:nifL protein, Bacteria; D005182:Flavin-Adenine Dinucleotide; D000641:Ammonia; D007501:Iron; D010544:Peroxidases