title: Studies of the heme coordination and ligand binding properties of soluble guanylyl cyclase (sGC): characterization of Fe(II)sGC and Fe(II)sGC(CO) by electronic absorption and magnetic circular dichroism spectroscopies and failure of CO to activate the enzyme.
abstract: The mechanism of activation of soluble guanylyl cyclase by NO is poorly understood although it is clear that NO interacts with a heme group in the protein via formation of a heme-nitrosyl adduct. The objective of this study is to investigate the coordination environment of the heme in the enzyme spectroscopically in the presence of known heme ligands and to correlate the spectral characteristics with other heme proteins of known structure. Comparison of the electronic and magnetic circular dichroism (MCD) spectra for ferrous bovine soluble guanylyl cyclase (Fe(II)sGC) in the absence and presence of the common heme ligand CO with those of other hemoproteins suggests that histidine is an axial ligand to the heme iron in Fe(II)sGC. Further analysis indicates that Fe(II)sGC is predominantly bis-histidine ligated; the ratio of MCD signal intensity in the visible region to that in the Soret region is most consistent with an admixture of pentacoordinate and hexacoordinate ferrous heme in Fe(II)sGC at pH 7.8. Spectral changes upon CO binding have been correlated with the activity of the enzyme to determine the relationship between coordination structure and activity. Although CO clearly binds to Fe(II)sGC to form a six-coordinate adduct, it fails to significantly activate the enzyme regardless of heme content or CO concentration. In contrast, the extent of activation of sGC by NO is dependent on the heme content in the enzyme and on the concentration of NO. These observations are consistent with a mechanism for activation of soluble guanylyl cyclase in which the bond between the heme iron and the proximal histidine must be broken for activation to take place.
authors.affiliation: Department of Chemistry, University of Wisconsin, Madison 53706, USA.
authors.forename: J N
authors.initials: JN
authors.lastname: Burstyn
authors.forename: A E
authors.initials: AE
authors.lastname: Yu
authors.forename: E A
authors.initials: EA
authors.lastname: Dierks
authors.forename: B K
authors.initials: BK
authors.lastname: Hawkins
authors.forename: J H
authors.initials: JH
authors.lastname: Dawson
mesh_terms: D000818:Animals; D002248:Carbon Monoxide / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D002417:Cattle; D002942:Circular Dichroism; D004789:Enzyme Activation / Q000187:drug effects; D005296:Ferrous Compounds / Q000737:chemistry* / Q000378:metabolism; D005990:Glycerol / Q000494:pharmacology; D006162:Guanylate Cyclase / Q000737:chemistry* / Q000378:metabolism; D006418:Heme / Q000737:chemistry / Q000378:metabolism*; D006639:Histidine / Q000737:chemistry / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D008024:Ligands; D009569:Nitric Oxide / Q000494:pharmacology
chemical_list: D005296:Ferrous Compounds; D008024:Ligands; D009569:Nitric Oxide; D006418:Heme; D006639:Histidine; D002248:Carbon Monoxide; D006162:Guanylate Cyclase; D005990:Glycerol