The activity of the Escherichia coli transcription factor FNR is regulated by a change in oligomeric state.
The transcription factor FNR globally regulates gene expression in response to oxygen deprivation in Escherichia coli. To understand how oxygen deprivation activates FNR, a constitutively active FNR* mutant protein, DA154, was studied to determine how this mutant bypassed the normal regulation pathway. When purified from aerobically grown cells, the DA154 protein had a larger apparent native molecular mass and an increased affinity for a consensus FNR target site as compared with wild-type FNR prepared under identical conditions. These results suggested that FNR* DA154 may bypass the normal regulation pathway by converting FNR from an inactive monomer to an active dimer under aerobic conditions. To determine whether wild-type FNR is active as a dimer under anaerobic conditions, FNR mutants were isolated that inhibit the activity of wild-type FNR by forming mixed dimers (i.e., dominant-negative mutants). These dominant-negative FNR mutants were shown to have substitutions in the putative DNA-binding domain and to be defective in binding to a consensus FNR DNA target site in vitro. One representative dominant-negative mutant, EK209, was also shown to be unable to form mixed oligomers in vivo under aerobic conditions, suggesting that FNR may be monomeric in the inactive state. Taken together, these data have led us to propose that under anaerobic conditions FNR is a dimer that is active for DNA binding, and under aerobic conditions, FNR is inactivated by conversion to a monomer.
Lazazzera
B A
BA
Department of Bacteriology, University of Wisconsin, Madison 53706.
Bates
D M
DM
Kiley
P J
PJ
D000595:Amino Acid Sequence; D000693:Anaerobiosis; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000378:metabolism*; D001483:Base Sequence; D003545:Cysteine / Q000235:genetics; D004252:DNA Mutational Analysis; D004269:DNA, Bacterial / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins; D015964:Gene Expression Regulation, Bacterial; D005799:Genes, Dominant; D017930:Genes, Reporter; D007506:Iron-Sulfur Proteins; D008957:Models, Genetic; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D010641:Phenotype; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000096:biosynthesis; D017422:Sequence Analysis, DNA; D014157:Transcription Factors / Q000096:biosynthesis / Q000378:metabolism*
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C060704:FNR protein, E coli; D007506:Iron-Sulfur Proteins; D011994:Recombinant Proteins; D014157:Transcription Factors; D003545:Cysteine