Abstract
As one of the main receptors of a second messenger, cGMP, cGMP-dependent protein kinase (PKG) isoforms I and II regulate distinct physiological processes. The design of isoform-specific activators is thus of great biomedical importance and requires detailed structural information about PKG isoforms bound with activators, including accurate positions of hydrogen atoms and a description of the hydrogen bonding and water architecture. Here, we determined a 2.2 Å room-temperature joint X-ray/neutron (XN) structure of the human PKG II carboxyl cyclic nucleotide binding (CNB-B) domain bound with a potent PKG II activator, 8-pCPT-cGMP. The XN structure directly visualizes intermolecular interactions and reveals changes in hydrogen bonding patterns upon comparison to the X-ray structure determined at cryo-temperatures. Comparative analysis of the backbone hydrogen/deuterium exchange patterns in PKG II:8-pCPT-cGMP and previously reported PKG Iβ:cGMP XN structures suggests that the ability of these agonists to activate PKG is related to how effectively they quench dynamics of the cyclic nucleotide binding pocket and the surrounding regions.
| Original language | English |
|---|---|
| Pages (from-to) | 1833-1837 |
| Number of pages | 5 |
| Journal | Biochemistry |
| Volume | 57 |
| Issue number | 12 |
| DOIs | |
| State | Published - Mar 27 2018 |
Funding
*E-mail: [email protected]. Phone: +1 505 310 4184. *E-mail: [email protected]. Phone: +1 713 798 8411. ORCID Choel Kim: 0000-0002-3152-0020 Andrey Kovalevsky: 0000-0003-4459-9142 Author Contributions J.C.C., C.K., and A.K. designed the research. O.G., J.C.C., and A.K. performed the research. A.K. and M.P.B. collected neutron diffraction data. O.G. and A.K. performed the structural analysis. Funding O.G. and A.K. were supported by the U.S. Department of Energy’s (DOE) Office of Basic Energy Sciences. J.C.C. and C.K. were funded by National Institutes of Health Grant R01 GM090161. This work has been authored by UT-Battelle LLC under DOE Contract DE-AC05-00OR22725. Notes The authors declare no competing financial interest. The authors thank Liying Qin and Shelton Boyd for critical reading of the manuscript. The research at Oak Ridge National Laboratory’s High Flux Isotope Reactor (IMAGINE beamline) was sponsored by the Scientific User Facilities Division, Office of Basic Energy Sciences, U.S. Department of Energy. The authors thank Institut Laue Langevin (beamline LADI-III) for awarded neutron beam time.
| Funders | Funder number |
|---|---|
| Oak Ridge National Laboratory | |
| Office of Basic Energy Sciences | |
| Scientific User Facilities Division | |
| U.S. Department of Energy’s | |
| UT-Battelle LLC | |
| National Institutes of Health | R01 GM090161 |
| U.S. Department of Energy | DE-AC05-00OR22725 |
| National Institute of General Medical Sciences | R25GM056929 |
| Basic Energy Sciences |