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 beta1(1-194) and beta2(1-217) were compared to the full-length enzyme. The EPR spectrum of the beta2(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 beta1(1-194)-NO complex resembles the EPR spectrum of sGC in the absence of the activators. Low-temperature resonance Raman spectra of the beta1(1-194)-NO and beta2(1-217)-NO complexes show that the Fe-NO stretching vibration of the beta2(1-217)-NO complex (535 cm (-1)) is significantly different from that of the beta1(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.
Derbyshire
Emily R
ER
Department of Molecular and Cell Biology, University of California, Berkeley, California 94720-3320, USA.
Gunn
Alexander
A
Ibrahim
Mohammed
M
Spiro
Thomas G
TG
Britt
R David
RD
Marletta
Michael A
MA
D000494:Allosteric Regulation; D000818:Animals; D003198:Computer Simulation; D004578:Electron Spin Resonance Spectroscopy; D005296:Ferrous Compounds / Q000737:chemistry*; D006160:Guanosine Triphosphate / Q000737:chemistry; D006162:Guanylate Cyclase / Q000737:chemistry*; D006418:Heme / Q000737:chemistry; D009569:Nitric Oxide / Q000737:chemistry*; D051381:Rats; D012995:Solubility; D013059:Spectrum Analysis, Raman
D005296:Ferrous Compounds; D009569:Nitric Oxide; D006418:Heme; D006160:Guanosine Triphosphate; D006162:Guanylate Cyclase