CHARACTERIZATION OF FNR-ASTERISK MUTANT PROTEINS INDICATES 2 DISTINCTMECHANISMS FOR ALTERING OXYGEN REGULATION OF THE ESCHERICHIA-COLI TRANSCRIPTION FACTOR FNR

In order to gain insight into the mechanism by which the Escherichia c oli transcription factor FNR is activated in response to anaerobiosis, we have analyzed FNR mutant proteins which, unlike the wild-type pro tein, stimulate gene expression in the presence of oxygen in vivo. Cel l extracts containing seven different FNR mutant proteins were tested in vitro for the ability to bind to the FNR consensus DNA site in a g el retardation assay under aerobic conditions. At the concentration of protein tested, only extracts which contained FNR mutant proteins wi th amino acid substitutions at position 154 showed significant DNA bin ding, The three position-154 FNR mutant proteins could be further dis tinguished from the other mutant proteins by analysis of the in vivo p henotypes of FNR proteins containing amino acid substitutions at eith er of two essential cysteine residues. In the presence of oxygen, FNR mutant proteins with amino acid substitutions at position 154 were th e 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 pro teins 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 .