- Oral presentation
- Open Access
The crystal structures of PKG Iβ (92-227) with cGMP and cAMP reveal the molecular details of cyclic-nucleotide binding
© Kim et al; licensee BioMed Central Ltd. 2011
- Published: 1 August 2011
- Cyclic Nucleotide
- Isothermal Titration Calorimetry
- Cyclic Phosphate
- Isothermal Titration Calorimetry Experiment
- Cyclic Nucleotide Binding
Cyclic GMP is a crucial second messenger that translates extracellular signals into a variety of cellular responses. As a central mediator of the Nitric Oxide-cGMP signalling cascade, which regulates vascular tone, platelet aggregation, nociception and hipocampal/cerebellar learning, Cyclic GMP-dependent protein kinases (PKGs) represents an important drug target for treating hypertensive diseases and erectile dysfunction.
The fidelity of the NO-cGMP signalling pathway is largely dependent on PKG’s ability to selectively bind cGMP over cAMP. Although both cGMP and cAMP bind and activate PKG, cGMP preferentially activates PKG 60-100 fold better than cAMP; yet, little is known about the molecular features required for the cGMP selectivity of PKG. We have investigated the mechanism of cyclic nucleotide binding to PKG by determining crystal structures of the amino-terminal cyclic nucleotide-binding domain (CNBD-A) of human PKG I bound to either cGMP or cAMP. We also determined the structure of CNBD-A in the absence of bound nucleotide.
Data and refinement statistics
P6 2 22
P6 2 22
Cell constants (Å)
50 – 2.9
50 – 2.49
Overall B value( Å2)
Rmsd bond length (Å)
Rmsd bond angle(°)
Cyclic Nucleotides interacting with the cGMP pocket. Both cGMP and cAMP bond in the cGMP binding pocket are shown on the far left and right and their Isothermal titration calorimetry data binding to the PKG Iβ CNBD-A shown in the middle. The cGMP-binding site is marked with three different sites: the short P-helix together with conserved glutamate and arginine residues at the PBC which captures the sugar phosphate (Site 1); a key residue, Thr193 at the end of PBC that bridges the cyclic phosphate to the guanine ring (Site 2); and the β5-strand that provides a unique docking site for the guanine ring (Site 3). Unlike cGMP, cAMP binds in two different configurations, anti in one molecule (shown on the far right panel) and syn in the other with different sets of contacts. Although the sugar phosphates share the same set of contacts with the protein at site 1, each purine ring of cAMP shows different contacts with the protein at sites 2 and 3The calorimetric measurements for cAMP and of cGMP binding to PKG Iβ (92-227) were carried out using a VP-ITC calorimeter (MicroCal LLC, Northampton, MA).
Our findings suggest that CNBD-A binds cGMP in the syn conformation through its interaction with Thr193 and an unusual cis-peptide forming residues Leu172 and Cys173. Although these studies provide the first structural insights into cyclic nucleotide binding to PKG, our ITC results show only a two-fold preference for cGMP, indicating that other domains are required for the previously reported cyclic nucleotide selectivity.
- Kim JJ, Casteel DE, Huang G, Kwon TH, Ren RK, Zwart P, Headd JJ, Brown NG, Chow DC, Palzkill T, Kim C: The crystal structures of PKG Iβ (92-227) with cGMP and cAMP reveal the molecular details of cyclic-nucleotide binding. Manuscript in preparation.Google Scholar
This article is published under license to BioMed Central Ltd. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.