Cra-dependent transcriptional activation of the icd gene of Escherichia coli

The icd gene of Escherichia coli, encoding isocitrate dehydrogenase, was sh own to be expressed from two different promoters: the previously identified icd P1 and a newly detected second promoter, icd P2, whose expression is p ositively regulated by the catabolite repressor-activator protein Cra, form erly called FruR, In each case, we determined the mRNA start site by primer extension analysis of in vivo transcripts and examined the interaction of the icd control region with either RNA polymerase or Cra, We observed that (i) the Cra factor binds to and activates transcription from a site centere d at position -76.5 within the led P2 promoter region and (ii) three partic ular mutations in the C-terminal end of the alpha subunit of RNA polymerase (L262A, R265A, and N268A) considerably diminish transcription initiating f rom the icd P2 promoter, as shown by in vitro experiments performed in the presence of mutant RNA polymerases carrying Ala substitutions.