Stringent spacing requirements for transcription activation by CRP

The cyclic AMP receptor protein-cAMP complex (CRP-cAMP) binds at a variety of distances upstream of several E. coli promoters and activates transcription. We have constructed a model system in which a consensus CRP binding site is placed at different distances upstream of the melR promoter. CRP-cAMP activates transcription from melR when bound at a number of positions, all of which lie on the same face of the DNA helix. The two distances at which transcription is strongly activated correspond exactly to those at which CRP-cAMP binds upstream of the well-studied galP1 and lac promoters. Footprinting of the synthetic promoters reveals that RNA polymerase makes identical contacts with their -10 regions even though CRP-cAMP binds at a different distance in each case. Kinetic analysis in vitro indicates that CRP-cAMP activates transcription from these promoters in similar but distinct ways. A model is proposed to explain this two-position activation.