Structural understanding of the allosteric conformational change of cyclicAMP receptor protein by cyclic AMP binding

Cyclic AMP receptor protein (CRP) plays a key role in the regulation of mor e than 150 genes. CRP is allosterically activated by cyclic AMP and binds t o specific DNA sites. A structural understanding of this allosteric conform ational change, which is essential for its function, is still lacking becau se the structure of apo-CRP has not been solved. Therefore, we performed va rious NMR experiments to obtain apo-CRP structural data. The secondary stru cture of apo-CRP was determined by analyses of the NOE connectivities, the amide proton exchange rates, and the H-1-N-15 steady-state NOE values. A co mbination of the CSI-method and TALOS prediction was also used to supplemen t the determination of the secondary structure of apo-CRP. This secondary s tructure of apo-CRP was compared with the known structure of cyclic AMP-bou nd CRP. The results suggest that the allosteric conformational change of CR P caused by cyclic AMP binding involves subunit realignment and domain rear rangement, resulting in the exposure of helix F onto the surface of the pro tein. Additionally, the results of the one-dimensional [C-13]carbonyl NMR e xperiments show that the conformational change of CRP caused by the binding of cyclic GMP, an analogue of cyclic AMP, is different from that caused by cyclic AMP binding.