Oxidation of cysteine 117 stimulates constitutive activation of the type Iα cGMP-dependent protein kinase.

Autor: Sheehe JL; From the Department of Pharmacology, Larner College of Medicine, and., Bonev AD; From the Department of Pharmacology, Larner College of Medicine, and., Schmoker AM; the Department of Biology, University of Vermont, Burlington, Vermont 05405 and., Ballif BA; the Department of Biology, University of Vermont, Burlington, Vermont 05405 and., Nelson MT; From the Department of Pharmacology, Larner College of Medicine, and., Moon TM; the Department of Chemistry and Biochemistry, University of Arizona, Tucson, Arizona 85721 thomasmoon@email.arizona.edu., Dostmann WR; From the Department of Pharmacology, Larner College of Medicine, and wolfgang.dostmann@uvm.edu.
Jazyk: angličtina
Zdroj: The Journal of biological chemistry [J Biol Chem] 2018 Oct 26; Vol. 293 (43), pp. 16791-16802. Date of Electronic Publication: 2018 Sep 11.
DOI: 10.1074/jbc.RA118.004363
Abstrakt: The type I cGMP-dependent protein kinase (PKG I) is an essential regulator of vascular tone. It has been demonstrated that the type Iα isoform can be constitutively activated by oxidizing conditions. However, the amino acid residues implicated in this phenomenon are not fully elucidated. To investigate the molecular basis for this mechanism, we studied the effects of oxidation using recombinant WT, truncated, and mutant constructs of PKG I. Using an in vitro assay, we observed that oxidation with hydrogen peroxide (H 2 O 2 ) resulted in constitutive, cGMP-independent activation of PKG Iα. PKG Iα C42S and a truncation construct that does not contain Cys-42 (Δ53) were both constitutively activated by H 2 O 2 In contrast, oxidation of PKG Iα C117S maintained its cGMP-dependent activation characteristics, although oxidized PKG Iα C195S did not. To corroborate these results, we also tested the effects of our constructs on the PKG Iα-specific substrate, the large conductance potassium channel (K Ca 1.1). Application of WT PKG Iα activated by either cGMP or H 2 O 2 increased the open probabilities of the channel. Neither cGMP nor H 2 O 2 activation of PKG Iα C42S significantly increased channel open probabilities. Moreover, cGMP-stimulated PKG Iα C117S increased K Ca 1.1 activity, but this effect was not observed under oxidizing conditions. Finally, we observed that PKG Iα C42S caused channel flickers, indicating dramatically altered K Ca 1.1 channel characteristics compared with channels exposed to WT PKG Iα. Cumulatively, these results indicate that constitutive activation of PKG Iα proceeds through oxidation of Cys-117 and further suggest that the formation of a sulfur acid is necessary for this phenotype.
(© 2018 Sheehe et al.)
Databáze: MEDLINE
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