Frank Wunder

Publications of Frank Wunder

• Chemosensory Ca2+ dynamics correlate with diverse behavioral phenotypes in human sperm.

  Authors: Thomas Veitinger, Jeffrey R Riffell, Sophie Veitinger, Jaclyn M Nascimento, Annika Triller, Charlie Chandsawangbhuwana, Katlen Schwane, Andreas Geerts, Frank Wunder, Michael W Berns, Eva M Neuhaus, Richard K Zimmer, Marc Spehr, Hanns Hatt

  The Journal of biological chemistry. 03/2011; 286(19):17311-25.

  In the female reproductive tract, mammalian sperm undergo a regulated sequence of prefusion changes that "prime" sperm for fertilization. Among the least understood of these complex processes are theIn the female reproductive tract, mammalian sperm undergo a regulated sequence of prefusion changes that "prime" sperm for fertilization. Among the least understood of these complex processes are the molecular mechanisms that underlie sperm guidance by environmental chemical cues. A "hard-wired" Ca(2+) signaling strategy that orchestrates specific motility patterns according to given functional requirements is an emerging concept for regulation of sperm swimming behavior. The molecular players involved, the spatiotemporal characteristics of such motility-associated Ca(2+) dynamics, and the relation between a distinct Ca(2+) signaling pattern and a behavioral sperm phenotype, however, remain largely unclear. Here, we report the functional characterization of two human sperm chemoreceptors. Using complementary molecular, physiological, and behavioral approaches, we comparatively describe sperm Ca(2+) responses to specific agonists of these novel receptors and bourgeonal, a known sperm chemoattractant. We further show that individual receptor activation induces specific Ca(2+) signaling patterns with unique spatiotemporal dynamics. These distinct Ca(2+) dynamics are correlated to a set of stimulus-specific stereotyped behavioral responses that could play vital roles during various stages of prefusion sperm-egg chemical communication.

• Discovery of Riociguat (BAY 63-2521): A Potent, Oral Stimulator of Soluble Guanylate Cyclase for the Treatment of Pulmonary Hypertension.

  Authors: Joachim Mittendorf, Stefan Weigand, Cristina Alonso-Alija, Erwin Bischoff, Achim Feurer, Michael Gerisch, Armin Kern, Andreas Knorr, Dieter Lang, Klaus Muenter, Martin Radtke, Hartmut Schirok, Karl-Heinz Schlemmer, Elke Stahl, Alexander Straub, Frank Wunder, Johannes-Peter Stasch

  ChemMedChem. 04/2009;

  Soluble guanylate cyclase (sGC) is a key signal-transduction enzyme activated by nitric oxide (NO). Impairments of the NO-sGC signaling pathway have been implicated in the pathogenesis ofSoluble guanylate cyclase (sGC) is a key signal-transduction enzyme activated by nitric oxide (NO). Impairments of the NO-sGC signaling pathway have been implicated in the pathogenesis of cardiovascular and other diseases. Direct stimulation of sGC represents a promising therapeutic strategy particularly for the treatment of pulmonary hypertension (PH), a disabling disease associated with a poor prognosis. Previous sGC stimulators such as the pyrazolopyridines BAY 41-2272 and BAY 41-8543 demonstrated beneficial effects in experimental models of PH, but were associated with unfavorable drug metabolism and pharmacokinetic (DMPK) properties. Herein we disclose an extended SAR exploration of this compound class to address these issues. Our efforts led to the identification of the potent sGC stimulator riociguat, which exhibits an improved DMPK profile and exerts strong effects on pulmonary hemodynamics and exercise capacity in patients with PH. Riociguat is currently being investigated in phase III clinical trials for the oral treatment of PH.

• Pharmacological characterization of receptor guanylyl cyclase reporter cell lines

  Authors: Frank Wunder, Daniel Barufe, Annette Woermann, Markus Milde

  BMC Pharmacology. 01/2009;

• A Novel PDE2A Reporter Cell Line: Characterization of the Cellular Activity of PDE Inhibitors.

  Authors: Frank Wunder, Mark Jean Gnoth, Andreas Geerts, Daniel Barufe

  Molecular pharmaceutics. 01/2009;

  We report here the generation and pharmacological characterization of a phosphodiesterase 2A (PDE2A) reporter cell line. Human PDE2A was stably transfected in a parental cell line expressing theWe report here the generation and pharmacological characterization of a phosphodiesterase 2A (PDE2A) reporter cell line. Human PDE2A was stably transfected in a parental cell line expressing the atrial natriuretic peptide (ANP) receptor and the cyclic nucleotide-gated (CNG) cation channel CNGA2, acting as the biosensor for intracellular cGMP. In this reporter cell line, cGMP levels can be monitored in real-time via aequorin luminescence stimulated by calcium influx through the CNG channel. By using different PDE inhibitors, we could show that our PDE2A reporter assay specifically monitors PDE2A inhibition with high sensitivity. In the absence of ANP stimulation, the PDE2A selective inhibitors EHNA, BAY 60-7550 and PDP did not increase basal luminescence levels in this experimental setting. However, in combination with ANP, these inhibitors stimulated luminescence signals and induced leftward shifts of ANP concentration-response curves. Similar results were obtained when using IBMX, trequinsin and dipyridamole, which inhibit PDE2A nonselectively with lower potency. PDP, the most potent PDE2A inhibitor known to date, was found to exhibit much lower cellular activity as anticipated from its biochemical PDE2A inhibitory activity. By cellular uptake and transport studies we could show that PDP's cell permeability is low and that the compound is a substrate for an efflux transporter. Other PDE inhibitors including vinpocetine, milrinone, rolipram, sildenafil, zaprinast, BRL 50481 and BAY 73-6691 did not stimulate luminescence signals on our PDE2A reporter cell line. The results imply that this novel PDE2A reporter assay provides an efficient, high throughput means for the identification and characterization of PDE2A inhibitors.

• Functional cell-based assays in microliter volumes for ultra-high throughput screening.

  Authors: Frank Wunder, Bernd Kalthof, Thomas Müller, Jörg Hüser

  Combinatorial chemistry & high throughput screening. 09/2008; 11(7):495-504.

  Functional cell-based assays have gained increasing importance for microplate-based high throughput screening (HTS). The use of high-density microplates, most prominently 1536-well plates, andFunctional cell-based assays have gained increasing importance for microplate-based high throughput screening (HTS). The use of high-density microplates, most prominently 1536-well plates, and miniaturized assay formats allow screening of comprehensive compound collections with more than 1 million compounds at ultra-high throughput, i.e. in excess of 100,000 samples per day. uHTS operations with numerous campaigns per year should generally support this throughput at all different steps of the process, including the underlying compound logistics, the (automated) testing of the corporate compound collection in the bioassay, and the subsequent follow-up studies for hit confirmation and characterization. A growing number of reports document the general feasibility of cell-based uHTS in microliter volumes. In addition, full automation with integrated robotic systems allows the realization of also complex assay protocols with multiple liquid handling and signal detection steps. For this review, cell-based assays are categorized based on the kinetics of the cellular response to be quantified in the test and the readout method employed. Thus, assays measuring fast cellular responses with high temporal resolution, e.g., receptor mediated calcium signals or changes in membrane potential, are at one end of this spectrum, while tests quantifying cellular transcriptional responses mark the opposite end. Trends for cell-based uHTS assays developed at Bayer-Schering Pharma are, first, to incorporate assay integral reference signals allowing the experimental differentiation of target hits from non-specifically acting compounds, and second, to make use of kinetic, real-time readouts providing additional information on the mode-of-action of test compounds.

• Pharmacological and kinetic characterization of adrenomedullin 1 and calcitonin gene-related peptide 1 receptor reporter cell lines.

  Authors: Frank Wunder, Annegret Rebmann, Andreas Geerts, Bernd Kalthof

  Molecular pharmacology. 05/2008; 73(4):1235-43.

  Adrenomedullin (ADM) and calcitonin gene-related peptide (CGRP) receptors and their respective ligands play important roles in cardiovascular (patho-)physiology. Functional expression of ADM and CGRPAdrenomedullin (ADM) and calcitonin gene-related peptide (CGRP) receptors and their respective ligands play important roles in cardiovascular (patho-)physiology. Functional expression of ADM and CGRP receptors requires the presence of the calcitonin receptor-like receptor (CRLR) together with receptor-activity-modifying proteins (RAMPs). We have characterized the expression patterns of CRLR and RAMP1 to RAMP3 in human cardiovascular-related tissues by quantitative polymerase chain reaction. We could identify high expression levels of CRLR, RAMP1, and RAMP2 in human heart and various blood vessels. RAMP3 expression in these tissues, however, was detectable at significantly lower levels. In addition, we describe here a novel, aequorin luminescence-based G protein-coupled receptor reporter assay that enables the real-time detection of receptor activation in living cells. In the assay system, intracellular cAMP levels are monitored with high sensitivity by using a modified, heteromultimeric cyclic nucleotide-gated channel mediating calcium influx. G(q)-coupled receptor activation is detected via aequorin luminescence stimulated by calcium release from intracellular stores. Using this novel reporter assay, we established and characterized stable ADM1 and CGRP1 receptor cell lines. The peptide ligands ADM, CGRP1, and CGRP2 were characterized as potent agonists at their respective receptors. In contrast, intermedin acted as a weak agonist on both receptors and showed only partial agonism on the ADM1 receptor. Agonist activities were effectively antagonized by the receptor antagonists ADM(22-52) and CGRP(8-37). Various vasoactive ADM fragments were also characterized but showed no activity on the ADM1 receptor cell line. In addition, luminescence signal kinetics after activation of G(s)- and G(q)-coupled receptors were found to be markedly different.

• Nitric oxide-independent activation of soluble guanylate cyclase by BAY 60-2770 in experimental liver fibrosis.

  Authors: Andreas Knorr, Claudia Hirth-Dietrich, Cristina Alonso-Alija, Michael Härter, Michael Hahn, Yvonne Keim, Frank Wunder, Johannes-Peter Stasch

  Arzneimittel-Forschung. 02/2008; 58(2):71-80.

  Liver cirrhosis is a chronic disease with high mortality rate and need for effective pharmacological intervention. The fibrotic remodelling of liver tissue is crucially dependent on hepatic stellateLiver cirrhosis is a chronic disease with high mortality rate and need for effective pharmacological intervention. The fibrotic remodelling of liver tissue is crucially dependent on hepatic stellate cell activation. Activation of hepatic stellate cells is reduced by an increase in cyclic guanosine monophosphate (cGMP). Stable cGMP analogues also reduce the contractile response of hepatic stellate cells. However, cGMP production is downregulated in the cirrhotic liver due to the reduced activity of the endothelial nitric oxide synthase. OBJECTIVE: Here we report that the novel activator of soluble guanylate cyclase (sGC), BAY 60-2770 (4-({(4-carboxybutyl) [2- (5-fluoro-2-{[4'-(trifluoromethyl) biphenyl-4-yl]methoxy}phenyl)ethyl] amino}methyl)benzoic acid), which increases the activity of sGC in a nitric oxide-independent manner, attenuates liver fibrosis in two rat models. METHODS: The compound was studied in the pig serum model and the carbon tetrachloride model. Fibrosis was assessed by estimating the increase in fibrous collagen by micromorphometry of histological sections stained with Sirius Red/Fast Green and by measuring total hepatic collagen. RESULTS: BAY 60-2770, on a recombinant sGC reporter cell line, stimulated the luminescence signals with an EC50 value of 5.4 +/- 1.2 nmol/L. In the presence of [1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ; 10 pmol/L) the EC50 was shifted to 0.39 +/- 0.11 nmol/L. In both fibrosis models, once daily oral administration of BAY 60-2770 concomittantly with the fibrotic stimulus prevented 60-75% of fibrosis, the lowest effective dose being 0.1 mg/kg in the pig serum model and 0.3 mg/kg in the carbon tetrachloride model. The treatment was well tolerated by all animals. The doses used were devoid of any significant influence on systemic blood pressure. Conclusion: Nitric oxide-independent activation of sGC might be an innovative therapeutic approach for the treatment of liver fibrosis of necro-inflammatory and immunological origin.

• A cell-based nitric oxide reporter assay useful for the identification and characterization of modulators of the nitric oxide/guanosine 3',5'-cyclic monophosphate pathway.

  Authors: Frank Wunder, Guido Buehler, Jörg Hüser, Stefan Mundt, Martin Bechem, Bernd Kalthof

  Analytical biochemistry. 05/2007; 363(2):219-27.

  Nitric oxide (NO) plays an important role in protection against the onset and progression of various cardiovascular disorders. Therefore, the NO/guanosine 3',5'-cyclic monophosphate (cGMP) pathwayNitric oxide (NO) plays an important role in protection against the onset and progression of various cardiovascular disorders. Therefore, the NO/guanosine 3',5'-cyclic monophosphate (cGMP) pathway has gained considerable attention and has become a target for new drug development. We have established a rapid, homogeneous, cell-based, and highly sensitive reporter assay for NO generated by endothelial nitric oxide synthase (eNOS). In a coculture system, NO production is indirectly monitored in living cells via soluble guanylyl cyclase (sGC) activation and calcium influx mediated by the olfactory cyclic nucleotide-gated (CNG) cation channel CNGA2, acting as the intracellular cGMP sensor. Using this NO reporter assay, we performed a fully automated high-throughput screening campaign for stimulators of NO synthesis. The coculture system reflects most aspects of the natural NO/cGMP pathway, namely, Ca(2+)-dependent and Ca(2+)-independent regulation of eNOS activity by G protein-coupled receptor agonists, oxidative stress, phosphorylation, and cofactor availability as well as NO-mediated stimulation of cGMP synthesis by sGC activation. The NO reporter assay allows the real-time detection of NO synthesis within living cells and makes it possible to identify and characterize activators and inhibitors of enzymes involved in the NO/cGMP signaling pathway.

• Design and synthesis of the first NO- and haem-independent sGC activator BAY 58–2667 for the treatment of acute decompensated heart failure

  Authors: Michael Hahn, Cristina Alonso-Alija, Friederike Stoll, Markus Heil, Joachim Mittendorf, Karl-Heinz Schlemmer, Frank Wunder, Johannes-Peter Stasch

  BMC Pharmacology. 01/2007;

• A luminescence-based assay for sensitive nitric oxide detection

  Authors: Frank Wunder, Guido Buehler, Jörg Hüser, Stefan Mundt, Martin Bechem, Bernd Kalthof

  BMC Pharmacology. 01/2007;

• Characterization of the first potent and selective PDE9 inhibitor using a cGMP reporter cell line.

  Authors: Frank Wunder, Adrian Tersteegen, Annegret Rebmann, Christina Erb, Thomas Fahrig, Martin Hendrix

  Molecular pharmacology. 01/2006; 68(6):1775-81.

  We report here the in vitro characterization of 1-(2-chlorophenyl)-6-[(2R)-3,3,3-trifluoro-2-methylpropyl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one (BAY 73-6691), the first potent and selectiveWe report here the in vitro characterization of 1-(2-chlorophenyl)-6-[(2R)-3,3,3-trifluoro-2-methylpropyl]-1,5-dihydro-4H-pyrazolo[3,4-d]pyrimidine-4-one (BAY 73-6691), the first potent and selective inhibitor of phosphodiesterase 9 (PDE9), which is currently under preclinical development for the treatment of Alzheimer's disease. This compound selectively inhibits human (IC50 = 55 nM) and murine (IC50 = 100 nM) PDE9 activity in vitro and shows only moderate activity against other cyclic nucleotide-specific phosphodiesterases. We also report the generation and characterization of a stably transfected PDE9 Chinese hamster ovary cell line, additionally expressing soluble guanylate cyclase (sGC), the olfactory cyclic nucleotide-gated cation channel CNGA2 and the photoprotein aequorin. In this cell line, intracellular cGMP levels can be monitored in real-time via aequorin luminescence induced by Ca2+ influx through CNGA2, acting as the intracellular cGMP sensor. This simple and sensitive assay system was used for the characterization of the cellular activity of the new PDE9 inhibitor. BAY 73-6691 alone did not significantly increase basal cGMP levels in this experimental setting. However, in combination with submaximal stimulating concentrations of the sGC activator 4-[((4-carboxybutyl)[2-[(4-phenethyl-benzyl)oxy]phenethyl]amino)methyl] benzoic acid (BAY 58-2667), the compound induced concentration-dependent luminescence signals and intracellular cGMP accumulation. The PDE9 inhibitor significantly potentiated the cGMP signals generated by sGC activating compounds such as BAY 58-2667 or 5-cyclopropyl-2-[1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-4-ylamine (BAY 41-2272) and induced leftward shifts of the corresponding concentration-response curves. Using our newly generated PDE9 reporter cell line, we could show that BAY 73-6691 is able to efficiently penetrate cells and to inhibit intracellular PDE9 activity.

• A cell-based cGMP assay useful for ultra-high-throughput screening and identification of modulators of the nitric oxide/cGMP pathway.

  Authors: Frank Wunder, Johannes-Peter Stasch, Joachim Hütter, Cristina Alonso-Alija, Jörg Hüser, Emanuel Lohrmann

  Analytical biochemistry. 05/2005; 339(1):104-12.

  We have established a rapid, homogeneous, cell-based, and highly sensitive assay for guanosine 3'-5'-cyclic monophosphate (cGMP) that is suitable for fully automated ultra-high-throughput screening.We have established a rapid, homogeneous, cell-based, and highly sensitive assay for guanosine 3'-5'-cyclic monophosphate (cGMP) that is suitable for fully automated ultra-high-throughput screening. In this assay system, cGMP production is monitored in living cells via Ca2+ influx through the olfactory cyclic nucleotide-gated cation channel CNGA2, acting as the intracellular cGMP sensor. A stably transfected Chinese hamster ovary (CHO) cell line was generated recombinantly expressing soluble guanylate cyclase, CNGA2, and aequorin as a luminescence indicator for the intracellular calcium concentration. This cell line was used to screen more than 900,000 compounds in an automated ultra-high-throughput screening assay using 1536-well microtiter plates. In this way, we have been able to identify BAY 58-2667, a member of a new class of amino dicarboxylic acids that directly activate soluble guanylate cyclase. The assay system allows the real-time cGMP detection within living cells and makes it possible to screen for activators and inhibitors of enzymes involved in the nitric oxide/cGMP pathway.

• Residues stabilizing the heme moiety of the nitric oxide sensor soluble guanylate cyclase.

  Authors: Peter M Schmidt, Christiane Rothkegel, Frank Wunder, Henning Schröder, Johannes-Peter Stasch

  European journal of pharmacology. 05/2005; 513(1-2):67-74.

  Soluble guanylate cyclase, a heterodimer consisting of an alpha- and a heme-containing beta-subunit, is the major receptor for the biological messenger nitric oxide (NO) and is involved in variousSoluble guanylate cyclase, a heterodimer consisting of an alpha- and a heme-containing beta-subunit, is the major receptor for the biological messenger nitric oxide (NO) and is involved in various signal transduction pathways. The heme moiety of the enzyme is bound between the axial heme ligand histidine(105) and the recently identified counterparts of the heme propionic acids, tyrosine(135) and arginine(139). The latter residues together with an invariant serine(137) form the unique heme binding motif Y-x-S-x-R. In this work, we show that replacement of the serine(137) with alanine destabilizes the binding of the heme moiety and impairs NO-mediated soluble guanylate cyclase activation.

• Beyond NO and heme: biochemical and pharmacological opportunities

  Authors: Christiane Rothkegel, Peter Schmidt, Arun HS, Friederike Stoll, Harald Lapp, Frank Wunder, Henning Schröder, Matthias Rinke, Harald Schmidt, Johannes-Peter Stasch

  BMC Pharmacology. 01/2005;

• Residues stabilizing the heme moiety of the NO sensor soluble guanylate cyclase

  Authors: Peter Schmidt, Christiane Rothkegel, Frank Wunder, Friederike Stoll, Henning Schröder, Johannes-Peter Stasch

  BMC Pharmacology. 01/2005;

• Characterization of the novel PDE9 inhibitor BAY 73-6691 using a cGMP reporter cell line

  Authors: Frank Wunder, Adrian Tersteegen, Martin Hendrix

  BMC Pharmacology. 01/2005;

• Antifibrotic effects of an sGC activator in rat models of liver fibrosis

  Authors: Claudia Hirth-Dietrich, Cristina Alonso-Alija, Michael Härter, Michael Hahn, Yvonne Keim, Frank Wunder, Andreas Knorr, Johannes-Peter Stasch

  BMC Pharmacology. 01/2005;

• NO- and haem-independent activation of soluble guanylyl cyclase: molecular basis and cardiovascular implications of a new pharmacological principle.

  Authors: Johannes-Peter Stasch, Peter Schmidt, Cristina Alonso-Alija, Heiner Apeler, Klaus Dembowsky, Michael Haerter, Markus Heil, Torsten Minuth, Elisabeth Perzborn, Ulrich Pleiss, Matthias Schramm, Werner Schroeder, Henning Schröder, Elke Stahl, Wolfram Steinke, Frank Wunder

  British journal of pharmacology. 08/2002; 136(5):773-83.

  1. Soluble guanylyl cyclase (sGC) is the only proven receptor for the ubiquitous biological messenger nitric oxide (NO) and is intimately involved in many signal transduction pathways, most notably1. Soluble guanylyl cyclase (sGC) is the only proven receptor for the ubiquitous biological messenger nitric oxide (NO) and is intimately involved in many signal transduction pathways, most notably in regulating vascular tone and platelet function. sGC is a heterodimeric (alpha/ss) protein that converts GTP to cyclic GMP; NO binds to its prosthetic haem group. Here, we report the discovery of a novel sGC activating compound, its interaction with a previously unrecognized regulatory site and its therapeutic implications. 2. Through a high-throughput screen we identified BAY 58-2667, an amino dicarboxylic acid which potently activates sGC in an NO-independent manner. In contrast to NO, YC-1 and BAY 41-2272, the sGC stimulators described recently, BAY 58-2667 activates the enzyme even after it has been oxidized by the sGC inhibitor ODQ or rendered haem deficient. 3. Binding studies with radiolabelled BAY 58-2667 show a high affinity site on the enzyme. 4. Using photoaffinity labelling studies we identified the amino acids 371 (alpha-subunit) and 231 - 310 (ss-subunit) as target regions for BAY 58-2667. 5. sGC activation by BAY 58-2667 results in an antiplatelet activity both in vitro and in vivo and a potent vasorelaxation which is not influenced by nitrate tolerance. 6. BAY 58-2667 shows a potent antihypertensive effect in conscious spontaneously hypertensive rats. In anaesthetized dogs the hemodynamic effects of BAY 58-2667 and GTN are very similar on the arterial and venous system. 7. This novel type of sGC activator is a valuable research tool and may offer a new approach for treating cardiovascular diseases.

• Heterogeneous Expression Patterns of Mammalian Potassium Channel Genes in Developing and Adult Rat Brain.

  Authors: Wilfried A. Kues, Frank Wunder

  The European journal of neuroscience. 02/1992; 4(12):1296-1308.

  Voltage-gated K+ channels in the mammalian brain are functionally heterogeneous. Mechanisms which may underlie heterogeneity are the expression of multiple K+ channel subunit genes, alternativeVoltage-gated K+ channels in the mammalian brain are functionally heterogeneous. Mechanisms which may underlie heterogeneity are the expression of multiple K+ channel subunit genes, alternative splicing and the formation of heteromultimers from different subunits. To examine the molecular basis of regional and cell-specific K+ channel expression in rat brain in situ hybridization techniques were used. The transcript distribution patterns of 11 cloned mammalian K+ channel genes encoding both slow- and fast-inactivating K+ channels from four different gene families were examined at different stages of development. The results show that each subunit-specific messenger RNA (mRNA) is independently expressed and is characterized by an individual expression pattern. In the hippocampal formation transcripts of RCK2, RCK3, RCK4, RCK5, Raw3 and rat Shal genes are heterogeneously expressed and regulated during postnatal development. RCK1, Raw1, Raw2 and DRK1 mRNAs, on the other hand, are present in the hippocampus throughout postnatal life. The expression patterns of the 11 genes partially overlap, suggesting the formation of different heteromultimeric K+ channel complexes.

• Residues stabilizing the heme moiety of the nitric oxide sensor soluble guanylate cyclase

  Authors: Peter M. Schmidt, Christiane Rothkegel, Frank Wunder, Henning Schröder, Johannes-Peter Stasch

  European Journal of Pharmacology.

  Soluble guanylate cyclase, a heterodimer consisting of an α- and a heme-containing β-subunit, is the major receptor for the biological messenger nitric oxide (NO) and is involved in various signalSoluble guanylate cyclase, a heterodimer consisting of an α- and a heme-containing β-subunit, is the major receptor for the biological messenger nitric oxide (NO) and is involved in various signal transduction pathways. The heme moiety of the enzyme is bound between the axial heme ligand histidine105 and the recently identified counterparts of the heme propionic acids, tyrosine135 and arginine139. The latter residues together with an invariant serine137 form the unique heme binding motif Y-x-S-x-R. In this work, we show that replacement of the serine137 with alanine destabilizes the binding of the heme moiety and impairs NO-mediated soluble guanylate cyclase activation.