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Neutron Crystallography Detects Differences in Protein Dynamics: Structure of the PKG II Cyclic Nucleotide Binding Domain in Complex with an Activator
journal contributionposted on 2018-03-08, 00:00 authored by Oksana Gerlits, James C. Campbell, Matthew P. Blakeley, Choel Kim, Andrey Kovalevsky
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.
Comparative analysisisoform-specific activatorsX-ray structurewater architectureCNB-BPKG isoformscyclic nucleotide binding pocketProtein Dynamics2.2 Å room-temperaturePKG II Cyclic Nucleotide Binding Domainhydrogen atomsNeutron Crystallography Detects DifferencesXN structurecGMP-dependent protein kinasePKG II carboxyl cyclic nucleotide bindingPKG II activator8- pCPT-cGMP