Christiane Rothkegel

Publications of Christiane Rothkegel

• Dimerization region of soluble guanylate cyclase characterized by bimolecular fluorescence complementation in vivo.

  Authors: Christiane Rothkegel, Peter M Schmidt, Derek-John Atkins, Linda Sarah Hoffmann, Harald H H W Schmidt, Henning Schröder, Johannes-Peter Stasch

  Molecular pharmacology. 12/2007; 72(5):1181-90.

  The ubiquitously expressed nitric oxide (NO) receptor soluble guanylate cyclase (sGC) plays a key role in signal transduction. Binding of NO to the N-terminal prosthetic heme moiety of sGC results inThe ubiquitously expressed nitric oxide (NO) receptor soluble guanylate cyclase (sGC) plays a key role in signal transduction. Binding of NO to the N-terminal prosthetic heme moiety of sGC results in approximately 200-fold activation of the enzyme and an increased conversion of GTP into the second messenger cGMP. sGC exists as a heterodimer the dimerization of which is mediated mainly by the central region of the enzyme. In the present work, we constructed deletion mutants within the predicted dimerization region of the sGC alpha(1)- and beta(1)-subunit to precisely map the sequence segments crucial for subunit dimerization. To track mutation-induced alterations of sGC dimerization, we used a bimolecular fluorescence complementation approach that allows visualizing sGC heterodimerization in a noninvasive manner in living cells. Our study suggests that segments spanning amino acids alpha(1)363-372, alpha(1)403-422, alpha(1)440-459, beta(1)212-222, beta(1)304-333, beta(1)344-363, and beta(1)381-400 within the predicted dimerization region are involved in the process of heterodimerization and therefore in the expression of functional sGC.

• Insights into activation and dimerization of soluble guanylate cyclase

  Authors: Christiane Rothkegel, Peter Schmidt, Derek-John Atkins, Friederike Stoll, Henning Schröder, Harald Schmidt, Johannes-Peter Stasch

  BMC Pharmacology. 01/2007;

• Targeting the heme-oxidized nitric oxide receptor for selective vasodilatation of diseased blood vessels.

  Authors: Johannes-Peter Stasch, Peter M Schmidt, Pavel I Nedvetsky, Tatiana Y Nedvetskaya, Arun Kumar H S, Sabine Meurer, Martin Deile, Ashraf Taye, Andreas Knorr, Harald Lapp, Helmut Müller, Yagmur Turgay, Christiane Rothkegel, Adrian Tersteegen, Barbara Kemp-Harper, Werner Müller-Esterl, Harald H H W Schmidt

  The Journal of clinical investigation. 10/2006; 116(9):2552-61.

  ROS are a risk factor of several cardiovascular disorders and interfere with NO/soluble guanylyl cyclase/cyclic GMP (NO/sGC/cGMP) signaling through scavenging of NO and formation of the strongROS are a risk factor of several cardiovascular disorders and interfere with NO/soluble guanylyl cyclase/cyclic GMP (NO/sGC/cGMP) signaling through scavenging of NO and formation of the strong oxidant peroxynitrite. Increased oxidative stress affects the heme-containing NO receptor sGC by both decreasing its expression levels and impairing NO-induced activation, making vasodilator therapy with NO donors less effective. Here we show in vivo that oxidative stress and related vascular disease states, including human diabetes mellitus, led to an sGC that was indistinguishable from the in vitro oxidized/heme-free enzyme. This sGC variant represents what we believe to be a novel cGMP signaling entity that is unresponsive to NO and prone to degradation. Whereas high-affinity ligands for the unoccupied heme pocket of sGC such as zinc-protoporphyrin IX and the novel NO-independent sGC activator 4-[((4-carboxybutyl){2-[(4-phenethylbenzyl)oxy]phenethyl}amino) methyl [benzoic]acid (BAY 58-2667) stabilized the enzyme, only the latter activated the NO-insensitive sGC variant. Importantly, in isolated cells, in blood vessels, and in vivo, BAY 58-2667 was more effective and potentiated under pathophysiological and oxidative stress conditions. This therapeutic principle preferentially dilates diseased versus normal blood vessels and may have far-reaching implications for the currently investigated clinical use of BAY 58-2667 as a unique diagnostic tool and highly innovative vascular therapy.

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• Identification of residues crucially involved in soluble guanylate cyclase activation.

  Authors: Christiane Rothkegel, Peter M Schmidt, Friederike Stoll, Henning Schröder, Harald H H W Schmidt, Johannes-Peter Stasch

  FEBS letters. 08/2006; 580(17):4205-13.

  The ubiquitous heterodimeric nitric oxide (NO) receptor soluble guanylate cyclase (sGC) plays a key role in various signal transduction pathways. Binding of NO takes place at the prosthetic hemeThe ubiquitous heterodimeric nitric oxide (NO) receptor soluble guanylate cyclase (sGC) plays a key role in various signal transduction pathways. Binding of NO takes place at the prosthetic heme moiety at the N-terminus of the beta(1)-subunit of sGC. The induced structural changes lead to an activation of the catalytic C-terminal domain of the enzyme and to an increased conversion of GTP into the second messenger cyclic GMP (cGMP). In the present work we selected and substituted different residues of the sGC heme-binding pocket based on a sGC homology model. The generated sGC variants were tested in a cGMP reporter cell for their effect on the enzyme activation by heme-dependent (NO, BAY 41-2272) stimulators and heme-independent (BAY 58-2667) activators. The use of these experimental tools allows the enzyme's heme content to be explored in a non-invasive manner. Asp(44), Asp(45) and Phe(74) of the beta(1)-subunit were identified as being crucially important for functional enzyme activation. beta(1)Asp(45) may serve as a switch between different conformational states of sGC and point to a possible mechanism of action of the heme dependent sGC stimulator BAY 41-2272. Furthermore, our data shows that the activation profile of beta(1)IIe(145) Tyr is unchanged compared to the native enzyme, suggesting that Tyr(145) does not confer the ability to distinguish between NO and O(2). In summary, the present work further elucidated intramolecular mechanisms underlying the NO- and BAY 41-2272-mediated sGC activation and raises questions regarding the postulated role of Tyr(145) for ligand discrimination.

• NO-independent activation of soluble guanylate cyclase prevents disease progression in rats with 5/6 nephrectomy.

  Authors: Philipp Kalk, Michael Godes, Katharina Relle, Christiane Rothkegel, Andreas Hucke, Johannes-Peter Stasch, Berthold Hocher

  British journal of pharmacology. 07/2006; 148(6):853-9.

  1. Chronic renal disease is associated with oxidative stress, reduced nitric oxide (NO) availability and soluble guanylate cyclase (sGC) dysfunction. Recently, we discovered BAY 58-2667, a compound1. Chronic renal disease is associated with oxidative stress, reduced nitric oxide (NO) availability and soluble guanylate cyclase (sGC) dysfunction. Recently, we discovered BAY 58-2667, a compound activating heme-deficient or oxidized sGC in a NO-independent manner. 2. We assessed potential of BAY 58-2667 in preventing cardiac and renal target organ damage in rats with 5/6 nephrectomy. 3. Male Wistar rats were allocated to three groups: 5/6 nephrectomy, 5/6 nephrectomy treated with BAY 58-2667 and sham operation. Study period was 18 weeks: blood pressure and creatinine clearance were assessed repeatedly. At study end blood samples were taken and hearts and kidneys harvested for histological studies. 4. BAY 58-2667 markedly lowered blood pressure in animals with 5/6 nephrectomy (untreated versus treated animals: 189+/-14 versus 146+/-11 mmHg, P<0.001). Left ventricular weight, cardiac myocyte diameter as well as cardiac arterial wall thickness significantly decreased in comparison to untreated animals with 5/6 nephrectomy. Natriuretic peptide plasma levels were also improved by BAY 58-2667. Kidney function and morphology as assessed by creatinine clearance, glomerulosclerosis, interstitial and perivascular fibrosis of intrarenal arteries were likewise significantly improved by BAY 58-2667. 5. This is the first study showing that BAY 58-2667 effectively lowers blood pressure, reduces left ventricular hypertrophy and slows renal disease progression in rats with 5/6 nephrectomy by targeting mainly oxidized sGC. Therefore, BAY 58-2667 represents a novel pharmacological principle with potential clinical value in treatment of chronic renal disease.

• 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;

• 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.