An N-terminally truncated form of cyclic GMP-dependent protein kinase Iα (PKG Iα) is monomeric, autoinhibited, and provides a model for activation

The type I cGMP-dependent protein kinases (PKG I) serve essential physiological functions, including smooth muscle relaxation, cardiac remodeling, and platelet aggregation. These enzymes form homodimers through their N-terminal dimerization domains, a feature implicated in regulating their cooperative activation. Previous investigations into the activation mechanisms of PKG I isoforms have been largely influenced by structures of the cAMP-dependent protein kinase (PKA). Here, we examined PKG Iα activation by cGMP and cAMP by engineering a monomeric form that lacks N-terminal residues 1-53 (Δ53). We found that the construct exists as a monomer as assessed by whole-protein MS, size-exclusion chromatography, and small-angle X-ray scattering (SAXS). Reconstruction of the SAXS 3D envelope indicates that Δ53 has a similar shape to the heterodimeric RIα-C complex of PKA. Moreover, we found that the Δ53 construct is autoinhibited in its cGMP-free state and can bind to and be activated by cGMP in a manner similar to full-length PKG Iα as assessed by surface plasmon resonance (SPR) spectroscopy. However, we found that the Δ53 variant does not exhibit cooperative activation, and its cyclic nucleotide selectivity is diminished. These findings support a model in which, despite structural similarities, PKG Iα activation is distinct from that of PKA, and its cooperativity is driven by in trans interactions between protomers.

Publikationsart
Zeitschriftenbeiträge (peer-reviewed)
Titel
An N-terminally truncated form of cyclic GMP-dependent protein kinase Iα (PKG Iα) is monomeric, autoinhibited, and provides a model for activation
Medien
Journal of Biological Chemistry
Heft
21
Band
293
Autoren
Thomas M. Moon, Jessica L. Sheehe, Praveena Nukareddy, Prof. Dr. Lydia Nausch , Jessica Wohlfahrt, Dwight E. Matthews, Donald K. Blumenthal, Wolfgang R. Dostmann
Seiten
7916-7929
Veröffentlichungsdatum
30.03.2018
Zitation
Moon, Thomas M.; Sheehe, Jessica L.; Nukareddy, Praveena; Nausch, Lydia; Wohlfahrt, Jessica; Matthews, Dwight E.; Blumenthal, Donald K.; Dostmann, Wolfgang R. (2018): An N-terminally truncated form of cyclic GMP-dependent protein kinase Iα (PKG Iα) is monomeric, autoinhibited, and provides a model for activation. Journal of Biological Chemistry 293 (21), S. 7916-7929. DOI: 10.1074/jbc.RA117.000647