Dissection of a surface-exposed portion of the cAMP-CRP complex that mediates transcription activation and repression

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Abstract
The Escherichia coli cAMP receptor protein (CRP) is essential for the activation and repression of transcription initiation at promoters in the CytR regulon. CRP performs these activities by making direct protein-protein interactions to the alpha-subunits of RNA polymerase and to the CytR regulator. Strikingly, it has been shown that amino acids of CRP that are critical for communication with the two partner proteins are located in close proximity on the surface of CRP. Here, we have dissected this surface in order to pinpoint the 'repression region' of CRP and to assess whether it overlaps with the characterized 'activating region'. Our results established that residues 12, 13, 17, 105, 108 and 110 are essential for the interaction with CytR and confirmed that 'activating region' 2 of CRP is made up of residues 19, 21 and 101. In the crystallographic structure of the CRP-DNA complex, the two sets of determinants are located immediately adjacent to each other forming a consecutive surface-exposed patch. The 'repression region' is chemically complementary to the characterized region on CytR that is essential for protein-protein communication to CRP. Moreover, the results provide insight into the mechanism by which CytR might prevent CRP-mediated transcription.