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

Autor:	Moon TM; Department of Pharmacology, Larner College of Medicine, University of Vermont, Burlington, Vermont 05405. Electronic address: thomasmoon@email.arizona.edu., Sheehe JL; Department of Pharmacology, Larner College of Medicine, University of Vermont, Burlington, Vermont 05405., Nukareddy P; Department of Chemistry, University of Vermont, Burlington, Vermont 05405., Nausch LW; Department of Pharmacology, Larner College of Medicine, University of Vermont, Burlington, Vermont 05405., Wohlfahrt J; Department of Pharmacology, Larner College of Medicine, University of Vermont, Burlington, Vermont 05405., Matthews DE; Department of Chemistry, University of Vermont, Burlington, Vermont 05405., Blumenthal DK; Department of Pharmacology and Toxicology, University of Utah, Salt Lake City, Utah 84112., Dostmann WR; Department of Pharmacology, Larner College of Medicine, University of Vermont, Burlington, Vermont 05405. Electronic address: wolfgang.dostmann@uvm.edu.
Jazyk:	angličtina
Zdroj:	The Journal of biological chemistry [J Biol Chem] 2018 May 25; Vol. 293 (21), pp. 7916-7929. Date of Electronic Publication: 2018 Mar 30.
DOI:	10.1074/jbc.RA117.000647
Abstrakt:	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. (© 2018 by The American Society for Biochemistry and Molecular Biology, Inc.)
Databáze:	MEDLINE
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