Binding to DNA protects Neisseria meningitidis fumarate and nitrate reductase regulator (FNR) from oxygen.
Here, we report the overexpression, purification, and characterization of the transcriptional activator fumarate and nitrate reductase regulator from the pathogenic bacterium Neisseria meningitidis (NmFNR). Like its homologue from Escherichia coli (EcFNR), NmFNR binds a 4Fe-4S cluster, which breaks down in the presence of oxygen to a 2Fe-2S cluster and subsequently to apo-FNR. The kinetics of NmFNR cluster disassembly in the presence of oxygen are 2-3x slower than those previously reported for wild-type EcFNR, but similar to constitutively active EcFNR* mutants, consistent with earlier work in which we reported that the activity of FNR-dependent promoters in N. meningitidis is only weakly inhibited by the presence of oxygen (Rock, J. D., Thomson, M. J., Read, R. C., and Moir, J. W. (2007) J. Bacteriol. 189, 1138-1144). NmFNR binds to DNA containing a consensus FNR box sequence, and this binding stabilizes the iron-sulfur cluster in the presence of oxygen. Partial degradation of the 4Fe-4S cluster to a 3Fe-4S occurs, and this form remains bound to the DNA. The 3Fe-4S cluster is converted spontaneously back to a 4Fe-4S cluster under subsequent anaerobic reducing conditions in the presence of ferrous iron. The finding that binding to DNA stabilizes FNR in the presence of oxygen such that it has a half-life of approximately 30 min on the DNA has implications for our appreciation of how oxygen switches off FNR activatable genes in vivo.
Edwards
James
J
Department of Biology (Area 10), University of York, Heslington, York YO10 5YW, United Kingdom.
Cole
Lindsay J
LJ
Green
Jasper B
JB
Thomson
Melanie J
MJ
Wood
A Jamie
AJ
Whittingham
Jean L
JL
Moir
James W B
JW
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism*; D007506:Iron-Sulfur Proteins / Q000235:genetics / Q000378:metabolism*; D009345:Neisseria meningitidis / Q000235:genetics / Q000378:metabolism*; D010100:Oxygen / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000502:physiology; D011485:Protein Binding / Q000502:physiology; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D007506:Iron-Sulfur Proteins; D014157:Transcription Factors; D010100:Oxygen