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Probing Domain Interactions in Soluble Guanylate Cyclase

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† ‡ Department of Molecular and Cell Biology, Department of Chemistry, and §Department of Plant and Microbial Biology, University of California, Berkeley, California 94720, United States
Department of Chemistry, University of Washington, Seattle, Washington 98195-1700, United States
⊥ @ California Institute for Quantitative Biosciences and @Division of Physical Biosciences, University of California, Berkeley, California 94720-3220, United States
*University of California, QB3 Institute, 570 Stanley Hall, Berkeley, CA 94720-3220. Phone: (510) 666-2763. Fax: (510) 666-2765. E-mail: [email protected]
Cite this: Biochemistry 2011, 50, 20, 4281–4290
Publication Date (Web):April 14, 2011
Copyright © 2011 American Chemical Society

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    Eukaryotic nitric oxide (NO) signaling involves modulation of cyclic GMP (cGMP) levels through activation of the soluble isoform of guanylate cyclase (sGC). sGC is a heterodimeric hemoprotein that contains a Heme–Nitric oxide and OXygen binding (H-NOX) domain, a Per/ARNT/Sim (PAS) domain, a coiled-coil (CC) domain, and a catalytic domain. To evaluate the role of these domains in regulating the ligand binding properties of the heme cofactor of NO-sensitive sGC, we constructed chimeras by swapping the rat β1 H-NOX domain with the homologous region of H-NOX domain-containing proteins from Thermoanaerobacter tengcongensis, Vibrio cholerae, and Caenorhabditis elegans (TtTar4H, VCA0720, and Gcy-33, respectively). Characterization of ligand binding by electronic absorption and resonance Raman spectroscopy indicates that the other rat sGC domains influence the bacterial and worm H-NOX domains. Analysis of cGMP production in these proteins reveals that the chimeras containing bacterial H-NOX domains exhibit guanylate cyclase activity, but this activity is not influenced by gaseous ligand binding to the heme cofactor. The rat–worm chimera containing the atypical sGC Gcy-33 H-NOX domain was weakly activated by NO, CO, and O2, suggesting that atypical guanylate cyclases and NO-sensitive guanylate cyclases have a common molecular mechanism for enzyme activation. To probe the influence of the other sGC domains on the mammalian sGC heme environment, we generated heme pocket mutants (Pro118Ala and Ile145Tyr) in the β1 H-NOX construct (residues 1–194), the β1 H-NOX-PAS-CC construct (residues 1–385), and the full-length α1β1 sGC heterodimer (β1 residues 1–619). Spectroscopic characterization of these proteins shows that interdomain communication modulates the coordination state of the heme–NO complex and the heme oxidation rate. Taken together, these findings have important implications for the allosteric mechanism of regulation within H-NOX domain-containing proteins.

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    Spectra of sGC mutants (Figure S1), activity of sGC α1β1 P118A (Figure S2), NO dissociation time courses of β1 I145Y mutants (Figure S3 and Table S1), and resonance Raman spectra of β1(1–194) mutants in the FeII-unligated state (Figure S4). This material is available free of charge via the Internet at

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    Cited By

    This article is cited by 15 publications.

    1. Ryu Makino, Shinsuke Yazawa, Hiroshi Hori, and Yoshitsugu Shiro . Interactions of Soluble Guanylate Cyclase with a P-Site Inhibitor: Effects of Gaseous Heme Ligands, Azide, and Allosteric Activators on the Binding of 2′-Deoxy-3′-GMP. Biochemistry 2012, 51 (46) , 9277-9289.
    2. Yilin Liu, James R. Kincaid. Resonance Raman studies of gas sensing heme proteins. Journal of Raman Spectroscopy 2021, 52 (12) , 2516-2535.
    3. Xin Wang, Chunlong Zhang, Qirui Chen, Zhaowu Ma, Hui Liu, Jiangrong Huang. Guanylate cyclases link serotoninergic signaling to modulate ethanol-induced food intake in C. elegans. Biochemical and Biophysical Research Communications 2021, 567 , 29-34.
    4. Pierre Moënne-Loccoz, Erik T. Yukl, Hirotoshi Matsumura. Mechanisms of Nitric Oxide Sensing and Detoxification by Bacterial Hemoproteins. 2018, 351-369.
    5. Jie Pan, Hong Yuan, Xiaoxue Zhang, Huijuan Zhang, Qiming Xu, Yajun Zhou, Li Tan, Shingo Nagawa, Zhong-Xian Huang, Xiangshi Tan. Probing the Molecular Mechanism of Human Soluble Guanylate Cyclase Activation by NO in vitro and in vivo. Scientific Reports 2017, 7 (1)
    6. Michelle C. Krzyzanowski, Sarah Woldemariam, Jordan F. Wood, Aditi H. Chaubey, Chantal Brueggemann, Alexander Bowitch, Mary Bethke, Noelle D. L’Etoile, Denise M. Ferkey, . Aversive Behavior in the Nematode C. elegans Is Modulated by cGMP and a Neuronal Gap Junction Network. PLOS Genetics 2016, 12 (7) , e1006153.
    7. Haoran Xu, Yuebin Zhang, Lei Chen, Yan Li, Chen Li, Li Liu, Takashi Ogura, Teizo Kitagawa, Zhengqiang Li. Entry of water into the distal heme pocket of soluble guanylate cyclase β1 H-NOX domain alters the ligated CO structure: a resonance Raman and in silico simulation study. RSC Advances 2016, 6 (49) , 43707-43714.
    8. Melody G. Campbell, Eric S. Underbakke, Clinton S. Potter, Bridget Carragher, Michael A. Marletta. Single-particle EM reveals the higher-order domain architecture of soluble guanylate cyclase. Proceedings of the National Academy of Sciences 2014, 111 (8) , 2960-2965.
    9. Markéta Martínková, Kenichi Kitanishi, Toru Shimizu. Heme-based Globin-coupled Oxygen Sensors: Linking Oxygen Binding to Functional Regulation of Diguanylate Cyclase, Histidine Kinase, and Methyl-accepting Chemotaxis. Journal of Biological Chemistry 2013, 288 (39) , 27702-27711.
    10. Nils Griebenow, Hartmut Schirok, Joachim Mittendorf, Alexander Straub, Markus Follmann, Johannes-Peter Stasch, Andreas Knorr, Karl-Heinz Schlemmer, Gorden Redlich. Identification of acidic heterocycle-substituted 1H-pyrazolo[3,4-b]pyridines as soluble guanylate cyclase stimulators. Bioorganic & Medicinal Chemistry Letters 2013, 23 (5) , 1197-1200.
    11. Laleh Alisaraie, Yangxin Fu, Jack A. Tuszynski. Dynamic Change of Heme Environment in Soluble Guanylate Cyclase and Complexation of NO‐Independent Drug Agents with H‐NOX Domain. Chemical Biology & Drug Design 2013, 81 (3) , 359-381.
    12. E.S. Underbakke, N.B. Surmeli, B.C. Smith, S.L. Wynia-Smith, M.A. Marletta. Nitric Oxide Signaling. 2013, 241-262.
    13. Thomas G. Spiro, Alexandra V. Soldatova, Gurusamy Balakrishnan. CO, NO and O2 as vibrational probes of heme protein interactions. Coordination Chemistry Reviews 2013, 257 (2) , 511-527.
    14. Nathaniel B. Fernhoff, Emily R. Derbyshire, Eric S. Underbakke, Michael A. Marletta. Heme-assisted S-Nitrosation Desensitizes Ferric Soluble Guanylate Cyclase to Nitric Oxide. Journal of Biological Chemistry 2012, 287 (51) , 43053-43062.
    15. A. N. Meints, J. G. Pemberton, J. P. Chang. Nitric Oxide and Guanylate Cyclase Signalling are Differentially Involved in Gonadotrophin (LH) Release Responses to Two Endogenous GnRHs from Goldfish Pituitary Cells. Journal of Neuroendocrinology 2012, 24 (8) , 1166-1181.

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