Bacillus cereus Fnr binds a [4Fe-4S] cluster and forms a ternary complex with ResD and PlcR.
BACKGROUND
Bacillus cereus is a facultative anaerobe that causes diarrheal disease in humans. Diarrheal syndrome may result from the secretion of various virulence factors including hemolysin BL and nonhemolytic enterotoxin Nhe. Expression of genes encoding Hbl and Nhe is regulated by the two redox systems, ResDE and Fnr, and the virulence regulator PlcR. B. cereus Fnr is a member of the Crp/Fnr family of iron-sulfur (Fe-S) proteins. Only its apo-form has so far been studied. A major goal in deciphering the Fnr-dependent regulation of enterotoxin genes is thus to obtain and characterize holoFnr.


RESULTS
Fnr has been subjected to in vitro Fe-S cluster reconstitution under anoxic conditions. UV-visible and EPR spectroscopic analyses together with the chemical estimation of the iron content indicated that Fnr binds one [4Fe-4S]2+ cluster per monomer. Atmospheric O2 causes disassembly of the Fe-S cluster, which exhibited a half-life of 15 min in air. Holo- and apoFnr have similar affinities for the nhe and hbl promoter regions, while holoFnr has a higher affinity for fnr promoter region than apoFnr. Both the apo- and holo-form of Fnr interact with ResD and PlcR to form a ternary complex.


CONCLUSIONS
Overall, this work shows that incorporation of the [4Fe-4S]2+ cluster is not required for DNA binding of Fnr to promoter regions of hbl and nhe enterotoxin genes or for the formation of a ternary complex with ResD and PlcR. This points to some new unusual properties of Fnr that may have physiological relevance in the redox regulation of enterotoxin gene regulation.
Université d'Avignon et des Pays de Vaucluse, UMR408, Sécurité et Qualité des Produits d'Origine Végétale, F-84000, Avignon, France.
Julia
J
Esbelin
Yves
Y
Jouanneau
Catherine
C
Duport
D001409:Bacillus cereus / Q000737:chemistry* / Q000378:metabolism*; D001426:Bacterial Proteins / Q000378:metabolism*; D004269:DNA, Bacterial / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D007501:Iron / Q000032:analysis; D007506:Iron-Sulfur Proteins / Q000378:metabolism*; D046912:Multiprotein Complexes / Q000737:chemistry*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D055503:Protein Multimerization; D013057:Spectrum Analysis; D015534:Trans-Activators / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D007506:Iron-Sulfur Proteins; D046912:Multiprotein Complexes; C099480:PlcR protein, Bacillus; D015534:Trans-Activators; D014157:Transcription Factors; D007501:Iron