Characterization of FNR* mutant proteins indicates two distinct mechanisms for altering oxygen regulation of the Escherichia coli transcription factor FNR.
In order to gain insight into the mechanism by which the Escherichia coli transcription factor FNR* is activated in response to anaerobiosis, we have analyzed FNR mutant proteins which, unlike the wild-type protein, stimulate gene expression in the presence of oxygen in vivo. Cell extracts containing seven different FNR* mutant proteins were tested in vitro for the ability to bind to the FNR consensus DNA site in a gel retardation assay under aerobic conditions. At the concentration of protein tested, only extracts which contained FNR* mutant proteins with amino acid substitutions at position 154 showed significant DNA binding. The three position-154 FNR* mutant proteins could be further distinguished from the other mutant proteins by analysis of the in vivo phenotypes of FNR* proteins containing amino acid substitutions at either of two essential cysteine residues. In the presence of oxygen, FNR* mutant proteins with amino acid substitutions at position 154 were the least affected when either Cys-23 or Cys-122 was substituted for Ser. On the basis of these in vivo and in vitro analyses, FNR* mutant proteins appear to segregate into at least two classes. Thus, it appears that each class of FNR* substitutions alters the normal pathway of FNR activation in response to oxygen deprivation by a different mechanism.
Bates
D M
DM
Department of Bacteriology, University of Wisconsin, Madison 53706, USA.
Lazazzera
B A
BA
Kiley
P J
PJ
D000332:Aerobiosis; D000595:Amino Acid Sequence; D000693:Anaerobiosis; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D003545:Cysteine / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics*; D029968:Escherichia coli Proteins; D015964:Gene Expression Regulation, Bacterial; D007506:Iron-Sulfur Proteins; D007763:Lac Operon / Q000235:genetics; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009154:Mutation; D010100:Oxygen / Q000494:pharmacology*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D012694:Serine / Q000235:genetics; D013329:Structure-Activity Relationship
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C060704:FNR protein, E coli; D007506:Iron-Sulfur Proteins; D012694:Serine; D003545:Cysteine; D010100:Oxygen