Interactions of wild-type and mutant AmpR of Citrobacter freundii with target DNA

Abstract

The AmpR transcriptional activator for the chromosomal ampC beta-lactamase gene of Citrobacter freundii was found to interact with an operator sequence located in the 5' half of the 38 bp region protected by AmpR in DNase I footprinting experiments. AmpR binding was associated with significant DNA bending of target DNA. A glycine to glutamic acid alteration at position 102 in AmpR converts AmpR into a transcriptional activator even in the absence of beta-lactam inducer. AmpRG102E interacted with the operator binding sequence and induced DNA bending. A glycine to lysine alteration at residue 102 completely abolished the ability of AmpR to transcriptionally affect the ampC promoter, i.e. to repress in the absence of beta-lactam inducer and induce in the presence of beta-lactam. Nevertheless, AmpRG102K could repress the oppositely orientated ampR promoter. AmpRG102K could also specifically interact with the operator but the resulting complex migrated faster in gel retardation experiments and no significant DNA bending was observed.