Characterization of two different five-coordinate soluble guanylate cyclase ferrous-nitrosyl complexes

Soluble guanylate cyclase (sGC), a hemoprotein, is the primary nitric oxide (NO) receptor in higher eukaryotes. The binding of NO to sGC leads to the formation of a five-coordinate ferrous-nitrosyl complex and a several hundred-fold increase in cGMP synthesis. NO activation of sGC is influenced by GTP and the allosteric activators YC-1 and BAY 41-2272. Electron paramagnetic resonance (EPR) spectroscopy shows that the spectrum of the sGC ferrous-nitrosyl complex shifts in the presence of YC-1, BAY 41-2272, or GTP in the presence of excess NO relative to the heme. These molecules shift the EPR signal from one characterized by g(1) = 2.083, g(2) = 2.036, and g(3) = 2.012 to a signal characterized by g(1) = 2.106, g(2) = 2.029, and g(3) = 2.010. The truncated heme domain constructs beta 1(1-194) and beta 2(1-217) were compared to the full-length enzyme. The EPR spectrum of the beta 2(1-217)-NO complex is characterized by g(1) = 2.106, g(2) = 2.025, and g(3) = 2.010, indicating the protein is a good model for the sGC-NO complex in the presence of the activators, while the spectrum of the beta 1(1-194)-NO complex resembles the EPR spectrum of sGC in the absence of the activators. Low-temperature resonance Raman spectra of the beta 1(1-194)-NO and beta 2(1-217)-NO complexes show that the Fe-NO stretching vibration of the beta 2(1-217)-NO complex (535 cm(-1)) is significantly different from that of the beta 1(1-194)-NO complex (527 cm(-1)). This shows that sGC can adopt different five-coordinate ferrous nitrosyl conformations and suggests that the Fe-NO conformation characterized by this unique EPR signal and Fe-NO stretching vibration represents a highly active sGC state.