Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:4.6.1.2 (guanylate cyclase)
8,497 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Chlorite dismutase (EC 1.13.11.49), an enzyme capable of reducing chlorite to chloride while producing molecular oxygen, has been characterized using EPR and optical spectroscopy. The EPR spectrum of GR-1 chlorite dismutase shows two different high-spin ferric heme species, which we have designated 'narrow' (gx,y,z = 6.24, 5.42, 2.00) and 'broad' (gz,y,x = 6.70, 5.02, 2.00). Spectroscopic evidence is presented for a proximal histidine co-ordinating the heme iron center of the enzyme. The UV/visible spectrum of the ferrous enzyme and EPR spectra of the ferric hydroxide and imidazole adducts are characteristic of a heme protein with an axial histidine co-ordinating the iron. Furthermore, the substrate analogs nitrite and hydrogen peroxide have been found to bind to ferric chlorite dismutase. EPR spectroscopy of the hydrogen peroxide adduct shows the loss of both high-spin and low-spin ferric signals and the appearance of a sharp radical signal. The NO adduct of the ferrous enzyme exhibits a low-spin EPR signal typical of a five-co-ordinate heme iron nitrosyl adduct. It seems that the bond between the proximal histidine and the iron is weak and can be broken upon binding of NO. The midpoint potential, Em(Fe3+/2+) = -23 mV, of chlorite dismutase is higher than for most heme enzymes. The spectroscopic features and redox properties of chlorite dismutase are more similar to the gas-sensing hemoproteins, such as guanylate cyclase and the globins, than to the heme enzymes.
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PMID:Spectroscopic characterization and ligand-binding properties of chlorite dismutase from the chlorate respiring bacterial strain GR-1. 1235 22

The heme-regulated eukaryotic initiation factor 2alpha (eIF2alpha) kinase (HRI), which is found primarily in reticulocytes, contains an N-terminal heme-binding domain (NT-HBD). Binding of NO to the heme iron of the NT-HBD of HRI activates its eIF2alpha kinase activity, thus inhibiting the initiation of translation in reticulocyte lysate. The EPR spectrum of the NO-bound NT-HBD showed several derivative-shaped lines around g = 2.00, which is one of the well-documented signature patterns of a six-coordinate NO complex with histidine as the axial ligand. This is in sharp contrast to that of another prototypical NO-sensor protein, soluble guanylate cyclase (sGC), in which the NO binding to the heme iron disrupts the iron-histidyl bond forming a five-coordinate NO. The NO-mediated activation of HRI is, therefore, not triggered by the cleavage of the iron-histidyl bond. As evidenced by the resonance Raman spectra, two inactive forms of HRI, the ferrous ligand-unbound and the CO-bound states of the NT-HBD, contain a six-coordinate complex as found for the NO complex, indicating that the replacement of the sixth ligand of the heme iron is not sufficient to trigger the activation of HRI. Because the configuration of liganded NO is different from that of liganded CO, we propose that specific interactions between liganded NO and surrounding amino acid residues, which would not be formed in the CO complex, are responsible for the NO-induced activation of HRI.
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PMID:NO-induced activation mechanism of the heme-regulated eIF2alpha kinase. 1243 Oct 98

The benzylindazole compound YC-1 has been shown to activate soluble guanylate cyclase by increasing the sensitivity toward NO and CO. Here we report the action of YC-1 on the coordination of CO- and NO-hemes in the enzyme and correlate the events with the activation of enzyme catalysis. A single YC-1-binding site on the heterodimeric enzyme was identified by equilibrium dialysis. To explore the affect of YC-1 on the NO-heme coordination, the six-coordinate NO complex of the enzyme was stabilized by dibromodeuteroheme substitution. Using the dibromodeuteroheme enzyme, YC-1 converted the six-coordinate NO-heme to a five-coordinate NO-heme with a characteristic EPR signal that differed from that in the absence of YC-1. These results revealed that YC-1 facilitated cleavage of the proximal His-iron bond and caused geometrical distortion of the five-coordinate NO-heme. Resonance Raman studies demonstrated the presence of two iron-CO stretch modes at 488 and 521 cm(-1) specific to the YC-1-bound CO complex of the native enzyme. Together with the infrared C-O stretching measurements, we assigned the 488-cm(-1) band to the iron-CO stretch of a six-coordinate CO-heme and the 521-cm(-1) band to the iron-CO stretch of a five-coordinate CO-heme. These results indicate that YC-1 stimulates enzyme activity by weakening or cleaving the proximal His-iron bond in the CO complex as well as the NO complex.
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PMID:YC-1 facilitates release of the proximal His residue in the NO and CO complexes of soluble guanylate cyclase. 1254 Aug 39

The binding of NO to the iron heme in guanylate cyclase and other heme proteins induces the cleavage of the proximal histidine bonded to the metal. In this study we assess by means of density functional theory (DFT) electronic structure calculations the role of H-bonding to histidine in the modulation of this effect. We have considered in the first place a model of the isolated active site coordinated with imidazole and imidazolate to mimic the effects of a very strong H-bond. We have also investigated four selected ferrous heme proteins with different proximal histidine environments: the O(2) sensing FixL, horseradish peroxidase C, and the alpha and beta subunits of human hemoglobin. Our results indicate that polarization and charge transfer effects associated with H-bonding to the proximal histidine play a fundamental role in the modulation of the NO trans effect in heme proteins. We also find computational evidence suggesting that protein structural constraints may affect significantly the cleavage of the Fe-His bond.
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PMID:Modulation of the NO trans effect in heme proteins: implications for the activation of soluble guanylate cyclase. 1264 10

Cytochrome c ' (cyt c ') is found in the periplasmic space of denitrifying bacteria where it is thought to mediate the transfer of NO between the nitrogen-cycle enzymes dissimilatory nitrite reductase and nitric oxide reductase. It contains a 5-coordinate (5c) His-ligated haem that shares spectroscopic and ligand-binding properties with the haem group in the sensory domain of soluble guanylate cyclase (sGC). The latter is an extremely important enzyme involved in the control of vasodilation and blood clotting. Curiously, the enzyme is activated up to 200-fold by the binding of NO to the haem, whereas the binding of CO gives rise to only a mild stimulation of activity. Through X-ray crystallography we have studied NO and CO binding to cyt c '. CO binds to the distal face to give a 6-coordinate (6c) adduct. By contrast, NO binding gives rise to a 5c adduct through the displacement of the proximal His, to give a novel and unexpected proximal binding mode for NO. These results are also supported by a range of spectroscopies. In the absence of a crystal structure for sGC we propose that cyt c ' provides a structural model for the haem domain of this enzyme and thereby helps to explain the differential effects of NO and CO on its activity.
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PMID:A two-faced molecule offers NO explanation: the proximal binding of nitric oxide to haem. 1277 55

Flash photolysis studies on the five-coordinate heme nitrosyl of Alcaligenes xylosoxidans cytochrome c' were carried out to investigate the ramifications of its proximal nitrosyl ligand on NO release. Delta absorbance spectra recorded 5 ms after photolysis indicate that approximately 5% of the photolyzed hemes are converted to a five-coordinate high spin ferrous state, revealing that reattachment of the endogenous His ligand is fast enough to trap some of the photolyzed heme. Analysis of NO rebinding suggests that the photolyzed ferrous protein is initially in a strained conformation, which relaxes on a millisecond time scale. The strained ferrous heme appears to contain a significantly labilized Fe-His bond, which allows direct second-order rebinding to the proximal face at high NO-concentrations. In contrast, the NO-binding properties of the relaxed conformation are similar to those previously observed in stopped-flow studies, which showed that a five-coordinate heme-nitrosyl is formed via a six-coordinate intermediate. The discovery of a rapid proximal His ligand reattachment to NO-dissociated heme reveals a novel "kinetic trap" mechanism for lowering the five-coordinate heme nitrosyl population in response to decreased ambient NO concentrations. Thus, NO dissociation from the five-coordinate heme nitrosyl, whether thermal or photochemical, is followed by rapid, and only slowly reversible, His reattachment which acts to kinetically trap the heme in its five-coordinate ferrous state. Because return to the five-coordinate heme nitrosyl requires two NO-dependent steps, the protein uses a kind of kinetic amplification of the thermodynamic dissociation that occurs in response to decreased NO concentrations. The implications of this "kinetic-trap" mechanism for NO release from soluble guanylate cyclase are discussed.
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PMID:A novel kinetic trap for NO release from cytochrome c': a possible mechanism for NO release from activated soluble guanylate cyclase. 1290 95

Soluble guanylate cyclase (sGC), a heterodimer consisting of alpha- and beta-subunit, is the key enzyme of the NO/cGMP signaling pathway. The heme moiety ligated to the beta-subunit via His(105) is crucial for the activation of the enzyme by NO. In addition to this NO binding capability, the heme status of the enzyme influences the activity of non-NO sGC activators and sGC inhibitors. Different sGC activity profiles were observed in the presence, absence, or the oxidized form of heme. Modulating the heme status is therefore crucial for the investigation of the mechanism of sGC activation. Here, we present a simple and reliable procedure for the removal of the heme moiety of sGC that is capable of eliminating any traces of unbound heme and detergent from the sample mixture in one single step. Samples containing 15 microg sGC and the non-ionic detergent Tween 20 (2%) were incubated at 37 degrees C for 10 min and loaded onto centrifugal ion exchange columns. After centrifugation, heme was bound entirely to the ion exchanger and could not be eluted, even after incubation with 1M NaCl. Tween 20 was found completely within the flowthrough. Heme-free sGC was eluted from the ion exchanger after application of 300 mM NaCl. The absence of the heme moiety was confirmed by UV/Vis spectra and determination of the enzymatic activity. In summary, the described procedure is suitable for the preparation of very small amounts of highly purified heme-free sGC for the investigation of the mechanism of action of different types of sGC activators.
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PMID:Preparation of heme-free soluble guanylate cyclase. 1296 39

Soluble guanylate cyclase (sGC), a heterodimeric hemeprotein, is the only receptor for the biological messenger nitric oxide (NO) identified to date and is intimately involved in various signal transduction pathways. By using the recently discovered NO- and heme-independent sGC activator BAY 58-2667 and a novel cGMP reporter cell, we could distinguish between heme-containing and heme-free sGC in an intact cellular system. Using these novel tools, we identified the invariant amino acids tyrosine 135 and arginine 139 of the beta(1)-subunit as crucially important for both the binding of the heme moiety and the activation of sGC by BAY 58-2667. The heme is displaced by BAY 58-2667 due to a competition between the carboxylic groups of this compound and the heme propionic acids for the identified residues tyrosine 135 and arginine 139. This displacement results in the release of the axial heme ligand histidine 105 and to the observed activation of sGC. Based on these findings we postulate a signal transmission triad composed of histidine 105, tyrosine 135, and arginine 139 responsible for the enzyme activation by this compound and probably also for transducing changes in heme status and porphyrin geometry upon NO binding into alterations of sGC catalytic activity.
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PMID:Identification of residues crucially involved in the binding of the heme moiety of soluble guanylate cyclase. 1457 Aug 94

The regulation of cGMP levels is central to the normal process of phototransduction in both cone and rod photoreceptor cells. Two of the proteins involved in this process are the enzyme, retinal guanylate cyclase (retGC), and its activating protein (GCAP) through which activity is regulated via changes in cellular Ca2+ levels. Dominant cone-rod dystrophies arising from changes in retGC1 are essentially restricted to mutations in codon 838 and result in the replacement of a conserved arginine residue with either cysteine, histidine or serine. In all three cases, the effect of the substitution on the in vitro cyclase activity is a loss of Ca2+ sensitivity arising from an increased stability of the coiled-coil domain of the protein dimer and retention of cyclase activity. In contrast, mutations in the Ca2+-coordinating EF hands of GCAP1 result in dominant cone dystrophy; the consequences of these mutations is a reduced ability of the mutant protein to regulate retGC activity in response to changes in Ca2+ levels. Functionally therefore, the retGC2 and GCAP2 mutations are similar in reducing the feedback inhibition of Ca2+ on cyclase activity and thereby on cGMP levels in the photoreceptors.
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PMID:Dominant cone and cone-rod dystrophies: functional analysis of mutations in retGC1 and GCAP1. 1475 May 95

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.
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PMID:Resonance Raman study on synergistic activation of soluble guanylate cyclase by imidazole, YC-1 and GTP. 1513 28


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