Resonance Raman study on synergistic activation of soluble guanylate cyclase by imidazole, YC-1 and GTP.
Soluble guanylate cyclase (sGC), a physiological nitric oxide (NO) receptor, is a heme-containing protein and catalyzes the conversion of GTP to cyclic GMP. We found that 200 mM imidazole moderately activated sGC in the coexistence with 3-(5'-hydroxymethyl-2'-furyl)-1-benzylindazole (YC-1), although imidazole or YC-1 alone had little effect for activation. GTP facilitated this process. Resonance Raman spectra of imidazole complex of native sGC and CO-bound sGC (CO-sGC) have demonstrated that a simple heme adduct with imidazole at the sixth coordination position is not present for both sGC and CO-sGC below 200 mM of the imidazole concentration and that the Fe-CO stretching band (nuFe-CO)) appears at 492 cm(-1) in the presence of imidazole compared with 473 cm(-1) in its absence. Both frequencies fall on the line of His-coordinated heme proteins in the nuFe-CO vs nuC-O plot. However, it is stressed that the CO-heme of sGC becomes apparently photo-inert in a spinning cell in the presence of imidazole, suggesting the formation of five-coordinate CO-heme or of six-coordinate heme with a very weak trans ligand. These observations suggest that imidazole alters not only the polarity of heme pocket but also the coordination structure at the fifth coordination side presumably by perturbing the heme-protein interactions at propionic side chains. Despite the fact that the isolated sGC stays in the reduced state and is not oxidized by O(2), sGC under the high concentration of imidazole (1.2 M) yielded nu4 at 1373 cm(-1) even after its removal by gel-filtration, but addition of dithionite gave the strong nu4 band at 1360 cm(-1). This indicated that imidazole caused autoxidation of sGC.
Pal
Biswajit
B
Center for Integrative Bioscience, Okazaki National Research Institutes, Higashiyama 5-1, Myodaiji, Okazaki 444-8585, Japan.
Li
Zhengqiang
Z
Ohta
Takehiro
T
Takenaka
Shigeo
S
Tsuyama
Shingo
S
Kitagawa
Teizo
T
D000818:Animals; D002417:Cattle; D004357:Drug Synergism; D004789:Enzyme Activation / Q000187:drug effects; D006160:Guanosine Triphosphate / Q000008:administration & dosage / Q000378:metabolism / Q000494:pharmacology*; D006162:Guanylate Cyclase / Q000737:chemistry* / Q000378:metabolism*; D006418:Heme / Q000737:chemistry; D007093:Imidazoles / Q000008:administration & dosage / Q000494:pharmacology*; D066298:In Vitro Techniques; D007191:Indazoles / Q000008:administration & dosage / Q000494:pharmacology*; D012995:Solubility; D013059:Spectrum Analysis, Raman
D007093:Imidazoles; D007191:Indazoles; C090937:3-(5'-hydroxymethyl-2'-furyl)-1-benzylindazole; D006418:Heme; C029899:imidazole; D006160:Guanosine Triphosphate; D006162:Guanylate Cyclase