Instability in a coiled-coil signaling helix is conserved for signal transduction in soluble guanylyl cyclase

Andrzej Weichsel, Jessica A. Kievenaar, Roslyn Curry, Jacob T. Croft, William "Bill" Montfort

Research output: Contribution to journalArticle

4 Scopus citations

Abstract

How nitric oxide (NO) activates its primary receptor, α1/β1 soluble guanylyl cyclase (sGC or GC-1), remains unknown. Likewise, how stimulatory compounds enhance sGC activity is poorly understood, hampering development of new treatments for cardiovascular disease. NO binding to ferrous heme near the N-terminus in sGC activates cyclase activity near the C-terminus, yielding cGMP production and physiological response. CO binding can also stimulate sGC, but only weakly in the absence of stimulatory small-molecule compounds, which together lead to full activation. How ligand binding enhances catalysis, however, has yet to be discovered. Here, using a truncated version of sGC from Manduca sexta, we demonstrate that the central coiled-coil domain, the most highly conserved region of the ~150,000 Da protein, not only provides stability to the heterodimer but is also conformationally active in signal transduction. Sequence conservation in the coiled coil includes the expected heptad-repeating pattern for coiled-coil motifs, but also invariant positions that disfavor coiled-coil stability. Full-length coiled coil dampens CO affinity for heme, while shortening of the coiled coil leads to enhanced CO binding. Introducing double mutation αE447L/βE377L, predicted to replace two destabilizing glutamates with leucines, lowers CO binding affinity while increasing overall protein stability. Likewise, introduction of a disulfide bond into the coiled coil results in reduced CO affinity. Taken together, we demonstrate that the heme domain is greatly influenced by coiled-coil conformation, suggesting communication between heme and catalytic domains is through the coiled coil. Highly conserved structural imperfections in the coiled coil provide needed flexibility for signal transduction.

Original languageEnglish (US)
JournalProtein Science
DOIs
Publication statusAccepted/In press - Jan 1 2019

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Keywords

  • cGMP
  • GC-1
  • guanylate cyclase
  • leucine zipper
  • nitric oxide
  • stimulator compound
  • YC-1

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology

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