Characterization of residues involved in soluble guanylate cyclase activation

Antonia Marazioti, George Dalkas, George Spyroulias, Andreas Papapetropoulos

Research output: Contribution to journalArticlepeer-review

Abstract

Soluble guanylate cyclase (sGC) is a heterodimeric hemoprotein composed of two sub-units, α1 or α2 and β1 that plays a key role in various signal transduction pathways. The enzyme is stimulated by nitric oxide (NO) following its interaction with the heme moiety bound to the N-terminus of the β1 subunit. The aim of the present work was to identify residues in the regulatory domain of the β1 subunit that regulate the responsiveness of the enzyme on heme dependent and heme independent agonists. Molecular dynamics calculations suggested mutations to perturb the native network of electrostatic interactions at the area of Heme propionic groups. Respective mutants were generated by using site-directed mutagenesis. COS-7 cells were then co-transfected with the wild type α1 subunit and wild-type or mutated β1 subunit of sGC. The expression levels of the sGC subunits were confirmed by Western blot analysis. One of the mutants exhibited no responsiveness to neither NO nor HMR-1766, indicating that the generated enzyme was devoid of any activity. On the other hand, a residue was identified that was 45% less responsive to NO, when stimulated by sodium nitroprusside. Interestingly, cGMP generation of this latter mutant following activation by HMR-1766 was identical to that of wild-type sGC. In the present work we have identified residues that differentially affect the responsiveness to heme-NO-independent sGC activators versus NO-releasing stimulators.

Original languageEnglish (US)
Pages (from-to)56-57
Number of pages2
JournalEpitheorese Klinikes Farmakologias kai Farmakokinetikes
Volume27
Issue number1
StatePublished - May 27 2009
Externally publishedYes

Keywords

  • HMR-1766
  • Site-directed mutagenesis
  • SNP
  • Soluble guanylate cyclase

ASJC Scopus subject areas

  • Pharmacology (medical)
  • Pharmacology

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