Butyl isocyanide as a probe of the activation mechanism of soluble guanylate cyclase. Investigating the role of non-heme nitric oxide.
Nitric oxide (NO) is a physiologically relevant activator of the hemoprotein soluble guanylate cyclase (sGC). In the presence of NO, sGC is activated several hundredfold above the basal level by a mechanism that remains to be elucidated. The heme ligand n-butyl isocyanide (BIC) was used to probe the mechanism of NO activation of sGC. Electronic absorption spectroscopy was used to show that BIC binds to the sGC heme, forming a 6-coordinate complex with an absorbance maximum at 429 nm. BIC activates sGC 2-5-fold, and synergizes with the allosteric activator YC-1, to activate the enzyme 15-25-fold. YC-1 activates the sGC-BIC complex, and leads to an increase in both the V(max) and K(m). BIC was also used to probe the mechanism of NO activation. The activity of the sGC-BIC complex increases 15-fold in the presence of NO, without displacing BIC at the heme, which is consistent with previous reports that proposed the involvement of a non-heme NO binding site in the activation process.
Derbyshire
Emily R
ER
Department of Molecular, Lawrence Berkeley National Laboratory, University of California, Berkeley, CA 94720, USA.
Marletta
Michael A
MA
D000818:Animals; D001665:Binding Sites; D004789:Enzyme Activation; D006162:Guanylate Cyclase / Q000737:chemistry*; D006418:Heme / Q000737:chemistry*; D017953:Isocyanates / Q000737:chemistry*; D008024:Ligands; D009569:Nitric Oxide / Q000737:chemistry*; D046590:Spectroscopy, Electron Energy-Loss
D017953:Isocyanates; D008024:Ligands; D009569:Nitric Oxide; D006418:Heme; D006162:Guanylate Cyclase; C002509:butyl isocyanate