A molecular basis for nitric oxide sensing by soluble guanylate cyclase

Nitric oxide (NO) functions as a signaling agent by activation of the solub le isoform of guanylate cyclase (sGC), a heterodimeric hemoprotein. NO bind s to the heme of sGC and triggers formation of cGMP from GTP, Here we repor t direct kinetic measurements of the multistep binding of NO to sGC and cor relate these presteady state events with activation of enzyme catalysis. NO binds to sGC to form a six-coordinate, nonactivated, intermediate (k(on) > 1.4 x 10(8) M-1.s(-1) at 4 degrees C). Subsequent release of the axial his tidine heme ligand is shown to be the molecular step responsible for activa tion of the enzyme. The rate at which this step proceeds also depends on NO concentration (k = 2.4 x 10(5) M-1.s(-1) at 4 degrees C), thus identifying a novel mode of regulation by NO. NO binding to the isolated heme domain o f sGC was also rapid (k = 7.1 +/- 2 x 10(8) M-1.s(-1) at 4 degrees C); howe ver, no intermediate was observed. The data show that sGC acts as an extrem ely fast, specific, and highly efficient trap for NO and that cleavage of t he iron-histidine bond provides the driving force for activation of sGC. In addition, the kinetic data indicate that transport or stabilization of NO is not necessary for effective signal transmission.