Allosteric control of promoter DNA bending by cyclic AMP receptor and cyclic AMP.
The structure of the cyclic AMP receptor-promoter complex in solution was studied in the range of 0.2-50 microM cAMP by measurements of the electric birefringence at 0.1 M salt using a lac promoter DNA with 121 bp and with the CAP binding site at its center. An excess of protein required for complete conversion of the promoter DNA into the specific complex seems to be partly due to nonspecific binding. The specific complex is associated with a decay time constant of 1.36 micros at 3 degrees C, a positive birefringence, and a permanent dipole moment demonstrated by pulse reversal. These attributes were observed at cAMP concentrations between 3 and 50 muM and are characteristic of the specific complex. Model calculations demonstrate that the DNA bending angle under these conditions is 92 degrees . The observed positive birefringence does not result from the combination of the calculated quasi-permanent dipole and the orientation of the helix axes alone but is due to coupling of translational and rotational diffusion. When the cAMP concentration is decreased below 3 microM, the positive birefringence turns to a negative one with a transition center at 1.5 microM. The transition is too narrow for a model with induction of the specific cyclic AMP receptor-promoter complex after binding of a single cAMP to the cyclic AMP receptor dimer but is consistent with induction of this complex after binding of two cAMP molecules. The cyclic AMP receptor-promoter complex is driven into its specific bent form in vitro in the range of cAMP concentrations corresponding to that required for gene regulation in vivo.
Porschke
Dietmar
D
Max Planck Institut für biophysikalische Chemie, 37077 Göttingen, Germany. dpoersc@gwdg.de
D000494:Allosteric Regulation; D001665:Binding Sites; D001718:Birefringence; D000242:Cyclic AMP / Q000737:chemistry*; D004247:DNA / Q000737:chemistry; D046911:Macromolecular Substances / Q000737:chemistry; D009690:Nucleic Acid Conformation; D011401:Promoter Regions, Genetic; D011487:Protein Conformation; D011953:Receptors, Cyclic AMP / Q000737:chemistry*
D046911:Macromolecular Substances; D011953:Receptors, Cyclic AMP; D004247:DNA; D000242:Cyclic AMP