[en] [en] BACKGROUND AND PURPOSE: Venomous animals express numerous Kunitz-type peptides. The mambaquaretin-1 (MQ1) peptide identified from the Dendroaspis angusticeps venom is the most selective antagonist of the arginine-vasopressin V2 receptor (V2R) and the only unique Kunitz-type peptide active on a GPCR. We aimed to exploit other mamba venoms to enlarge the V2R-Kunitz peptide family and gain insight into the MQ1 molecular mode of action.
EXPERIMENTAL APPROACH: We used a bio-guided screening assay to identify novel MQs and placed them phylogenetically. MQs were produced by solid-phase peptide synthesis and characterized in vitro by binding and functional tests and in vivo by diuresis measurement in rats.
KEY RESULTS: Eight additional MQs were identified with nanomolar affinities for the V2R, all antagonists. MQs form a new subgroup in the Kunitz family, close to the V2R non-active dendrotoxins and to two V2R-active cobra toxins. Sequence comparison between active and non-active V2R Kunitz peptides highlighted five positions, among which four are involved in V2R interaction and belong to the two large MQ1 loops. We finally determined that eight positions, part of these two loops, interact with the V2R. The variant MQ1-K39A showed a higher affinity for the hV2R, but not for the rat V2R.
CONCLUSIONS AND IMPLICATIONS: A new function and mode of action is associated with the Kunitz peptides. The number of MQ1 residues involved in V2R binding is large and may explain its absolute selectivity. MQ1-K39A represents the first step in the improvement of the MQ1 design from a medicinal perspective.
Disciplines :
Biochemistry, biophysics & molecular biology
Author, co-author :
Droctové, Laura; Département Médicaments et Technologies pour la Santé (DMTS), SIMoS, Université Paris Saclay, CEA, INRAE, Gif-sur-Yvette, France
Ciolek, Justyna; Département Médicaments et Technologies pour la Santé (DMTS), SIMoS, Université Paris Saclay, CEA, INRAE, Gif-sur-Yvette, France
Mendre, Christiane; Institut de Génomique Fonctionnelle, Université de Montpellier, CNRS, INSERM, Montpellier, France
Chorfa, Amélia; Institut de Génomique Fonctionnelle, Université de Montpellier, CNRS, INSERM, Montpellier, France
Huerta, Paola; Département Médicaments et Technologies pour la Santé (DMTS), SIMoS, Université Paris Saclay, CEA, INRAE, Gif-sur-Yvette, France
Carvalho, Chrystelle; Département Médicaments et Technologies pour la Santé (DMTS), SIMoS, Université Paris Saclay, CEA, INRAE, Gif-sur-Yvette, France
Gouin, Charlotte; Département Médicaments et Technologies pour la Santé (DMTS), SIMoS, Université Paris Saclay, CEA, INRAE, Gif-sur-Yvette, France
Lancien, Manon; Département Médicaments et Technologies pour la Santé (DMTS), SIMoS, Université Paris Saclay, CEA, INRAE, Gif-sur-Yvette, France
Stanajic-Petrovic, Goran; Département Médicaments et Technologies pour la Santé (DMTS), SIMoS, Université Paris Saclay, CEA, INRAE, Gif-sur-Yvette, France
Braco, Lorine; Département Médicaments et Technologies pour la Santé (DMTS), SIMoS, Université Paris Saclay, CEA, INRAE, Gif-sur-Yvette, France
Blanchet, Guillaume; Département Médicaments et Technologies pour la Santé (DMTS), SIMoS, Université Paris Saclay, CEA, INRAE, Gif-sur-Yvette, France
Upert, Gregory; Département Médicaments et Technologies pour la Santé (DMTS), SIMoS, Université Paris Saclay, CEA, INRAE, Gif-sur-Yvette, France
De Pauw, Gregory; Laboratory of Mass Spectrometry, MolSys Research Unit, University of Liège, Liège, Belgium
Barbe, Peggy; Département Médicaments et Technologies pour la Santé (DMTS), SIMoS, Université Paris Saclay, CEA, INRAE, Gif-sur-Yvette, France
Keck, Mathilde; Département Médicaments et Technologies pour la Santé (DMTS), SIMoS, Université Paris Saclay, CEA, INRAE, Gif-sur-Yvette, France
Mourier, Gilles; Département Médicaments et Technologies pour la Santé (DMTS), SIMoS, Université Paris Saclay, CEA, INRAE, Gif-sur-Yvette, France
Mouillac, Bernard; Institut de Génomique Fonctionnelle, Université de Montpellier, CNRS, INSERM, Montpellier, France
Denis, Servent ; Département Médicaments et Technologies pour la Santé (DMTS), SIMoS, Université Paris Saclay, CEA, INRAE, Gif-sur-Yvette, France
Rodríguez de la Vega, Ricardo C; Ecologie, Systematique Evolution, CNRS, AgroParisTech, Université Paris-Saclay, Orsay, France
Quinton, Loïc ; Université de Liège - ULiège > Département de chimie (sciences) > Chimie biologique
Gilles, Nicolas; Département Médicaments et Technologies pour la Santé (DMTS), SIMoS, Université Paris Saclay, CEA, INRAE, Gif-sur-Yvette, France
CEA - Commissariat à l'Énergie Atomique et aux Énergies Alternatives LNCC - Ligue Nationale Contre le Cancer
Funding text :
We thank Mathilde Gilles for her contribution in improving the quality of English language. This work was supported by the French Atomic and Alternative Energies and La Ligue contre le Cancer for financial support and for the Amelia Chorfa PhD programme funding.This work was supported by the French Atomic and Alternative Energies and for financial support and for the Amelia Chorfa PhD programme funding. La Ligue contre le Cancer
Ainsworth, S., Petras, D., Engmark, M., Süssmuth, R. D., Whiteley, G., Albulescu, L., Kazandjian, T. D., Wagstaff, S. C., Rowley, P., Wüster, W., Dorrestein, P. C., Arias, A. S., Gutiérrez, J. M., Harrison, R. A., Casewell, N. R., & Calvete, J. J. (2018). The medical threat of mamba envenoming in sub-Saharan Africa revealed by genus-wide analysis of venom composition, toxicity and antivenomics profiling of available antivenoms. Journal of Proteomics, 172, 173–189. https://doi.org/10.1016/j.jprot.2017.08.016
Andreev, Y., Kozlov, S. A., Koshelev, S. G., Ivanova, E. A., Monastyrnaya, M. M., Kozlovskaya, E. P., & Grishin, E. V. (2008). Analgesic compound from sea anemone Heteractis crispa is the first polypeptide inhibitor of vanilloid receptor 1 (TRPV1). The Journal of Biological Chemistry, 283, 23914–23921. https://doi.org/10.1074/jbc.M800776200
Blanchet, G., Alili, D., Protte, A., Upert, G., Gilles, N., Tepshi, L., Stura, E. A., Mourier, G., & Servent, D. (2017). Ancestral protein resurrection and engineering opportunities of the mamba aminergic toxins. Scientific Reports, 7, 1–12. https://doi.org/10.1038/s41598-017-02953-0
Blanchet, G., Collet, G., Mourier, G., Gilles, N., Fruchart-Gaillard, C., Marcon, E., & Servent, D. (2014). Polypharmacology profiles and phylogenetic analysis of three-finger toxins from mamba venom: Case of aminergic toxins. Biochimie, 103, 109–117. https://doi.org/10.1016/j.biochi.2014.04.009
Bohlen, C. J., Chesler, A. T., Sharif-Naeini, R., Medzihradszky, K. F., Zhou, S., King, D., Sánchez, E. E., Burlingame, A. L., Basbaum, A. I., & Julius, D. (2011). A heteromeric Texas coral snake toxin targets acid-sensing ion channels to produce pain. Nature, 479, 410–414. https://doi.org/10.1038/nature10607
Bourne, Y., Taylor, P., & Marchot, P. (1995). Acetylcholinesterase inhibition by fasciculin: Crystal structure of the complex. Cell, 83, 503–512. https://doi.org/10.1016/0092-8674(95)90128-0
Ciolek, J., Reinfrank, H., Quinton, L., Viengchareun, S., Stura, E. A., Vera, L., Sigismeau, S., Mouillac, B., Orcel, H., Peigneur, S., Tytgat, J., Droctové, L., Beau, F., Nevoux, J., Lombès, M., Mourier, G., de Pauw, E., Servent, D., Mendre, C., … Gilles, N. (2017). Green mamba peptide targets type-2 vasopressin receptor against polycystic kidney disease. Proceedings of the National Academy of Sciences of the United States of America, 114, 7154–7159. https://doi.org/10.1073/pnas.1620454114
Cornett, L. E., & Dorsa, D. M. (1986). Regulation of (3H) arginine8 vasopressin binding to the rat renal medulla by guanine nucleotides. Journal of Receptor Research, 6, 127–140. https://doi.org/10.3109/10799898609073928
Curtis, M. J., Alexander, S., Cirino, G., Docherty, J. R., George, C. H., Giembycz, M. A., Hoyer, D., Insel, P. A., Izzo, A. A., Ji, Y., MacEwan, D. J., Sobey, C. G., Stanford, S. C., Teixeira, M. M., Wonnacott, S., & Ahluwalia, A. (2018). Experimental design and analysis and their reporting II: Updated and simplified guidance for authors and peer reviewers. British Journal of Pharmacology, 175(7), 987–993. https://doi.org/10.1111/bph.14153
Droctové, L., Lancien, M., Tran, V. L., Susset, M., Jego, B., Theodoro, F., Kessler, P., Mourier, G., Robin, P., Diarra, S. S., Palea, S., Flahault, A., Chorfa, A., Corbani, M., Llorens-Cortes, C., Mouillac, B., Mendre, C., Pruvost, A., Servent, D., … Gilles, N. (2020). A snake toxin as a theranostic agent for the type 2 vasopressin receptor. Theranostics, 10, 11580–11594. https://doi.org/10.7150/thno.47485
Fruchart-Gaillard, C., Mourier, G., Blanchet, G., Vera, L., Gilles, N., Ménez, R., Marcon, E., Stura, E. A., & Servent, D. (2012). Engineering of three-finger fold toxins creates ligands with original pharmacological profiles for muscarinic and adrenergic receptors. PLoS ONE, 7, e39166. https://doi.org/10.1371/journal.pone.0039166
Gasparini, S., Danse, J. M., Lecoq, A., Pinkasfeld, S., Zinn-Justin, S., Young, L. C., de Medeiros, C. C. L., Rowan, E. G., Harvey, A. L., & Ménez, A. (1998). Delineation of the functional site of α-dendrotoxin: The functional topographies of dendrotoxins are different but share a conserved core with those of other Kv1 potassium channel-blocking toxins. The Journal of Biological Chemistry, 273, 25393–25403. https://doi.org/10.1074/jbc.273.39.25393
Harper, E., & Berger, A. (1967). On the size of the active site in proteases: I. Papain. Biochemical and Biophysical Research Communications, 27, 157–162.
Harvey, A. L. (2001). Twenty years of dendrotoxins. Toxicon, 39, 15–26. https://doi.org/10.1016/S0041-0101(00)00162-8
Huber, R., Kukla, D., Rühlmann, A., Epp, O., & Formanek, H. (1970). The basic trypsin inhibitor of bovine pancreas. I. Structure analysis and conformation of the polypeptide chain. Naturwissenschaften, 57, 389–392. https://doi.org/10.1007/BF00599976
Izzo, A. A., Teixeira, M., Alexander, S. P., Cirino, G., Docherty, J. R., George, C. H., Insel, P. A., Ji, Y., Kendall, D. A., Panattieri, R. A., Sobey, C. G., Stanford, S. C., Stefanska, B., Stephens, G., & Ahluwalia, A. (2020). A practical guide for transparent reporting of research on natural products in the British Journal of Pharmacology: Reproducibility of natural product research. British Journal of Pharmacology, 177(10), 2169–2178. https://doi.org/10.1111/bph.15054
Juul, K. V., Bichet, D. G., Nielsen, S., & Nørgaard, J. P. (2014). The physiological and pathophysiological functions of renal and extrarenal vasopressin V2 receptors. American Journal of Physiology. Renal Physiology, 306, F931–F940. https://doi.org/10.1152/ajprenal.00604.2013
Kawamura, K., Yamada, T., Kurihara, K., Tamada, T., Kuroki, R., Tanaka, I., Takahashi, H., & Niimura, N. (2011). X-ray and neutron protein crystallographic analysis of the trypsin-BPTI complex. Acta Crystallographica. Section D, Biological Crystallography, 67, 140–148. https://doi.org/10.1107/S0907444910053382
Kessler, P., Marchot, P., Silva, M., & Servent, D. (2017). The three-finger toxin fold: A multifunctional structural scaffold able to modulate cholinergic functions. Journal of Neurochemistry, 142(Suppl), 7–18. https://doi.org/10.1111/jnc.13975
Kunitz, M., & Northrop, J. H. (1936). Isolation from beef pancreas of crystalline trypsinogen, trypsin, a trypsin inhibitor, and an inhibitor-trypsin compound. The Journal of General Physiology, 19, 991–1007. https://doi.org/10.1085/jgp.19.6.991
Lilley, E., Stanford, S. C., Kendall, D. E., Alexander, S. P., Cirino, G., Docherty, J. R., George, C. H., Insel, P. A., Izzo, A. A., Ji, Y., Panettieri, R. A., Sobey, C. G., Stefanska, B., Stephens, G., Teixeira, M., & Ahluwalia, A. (2020). ARRIVE 2.0 and the British Journal of Pharmacology: Updated guidance for 2020. British Journal of Pharmacology, 177(16), 3611–3616. https://doi.org/10.1111/bph.15178
Maeda, S., Xu, J., N. Kadji, F. M., Clark, M. J., Zhao, J., Tsutsumi, N., Aoki, J., Sunahara, R. K., Inoue, A., Garcia, K. C., & Kobilka, B. K. (2020). Structure and selectivity engineering of the M1 muscarinic receptor toxin complex. Science, 369, 161–167. https://doi.org/10.1126/science.aax2517
Maïga, A., Mourier, G., Quinton, L., Rouget, C., Gales, C., Denis, C., Lluel, P., Sénard, J. M., Palea, S., Servent, D., & Gilles, N. (2012). G protein-coupled receptors, an unexploited animal toxin targets: Exploration of green mamba venom for novel drug candidates active against adrenoceptors. Toxicon, 59, 487–496. https://doi.org/10.1016/j.toxicon.2011.03.009
Otlewski, J., Jaskólski, M., Buczek, O., Cierpicki, T., Czapińska, H., Krowarsch, D., Smalas, A. O., Stachowiak, D., Szpineta, A., & Dadlez, M. (2001). Structure-function relationship of serine protease-protein inhibitor interaction. Acta Biochimica Polonica, 48, 419–428. https://doi.org/10.18388/abp.2001_3926
Percie du Sert, N., Hurst, V., Ahluwalia, A., Alam, S., Avey, M. T., Baker, M., Browne, W. J., Clark, A., Cuthill, I. C., Dirnagl, U., Emerson, M., Garner, P., Holgate, S. T., Howells, D. W., Karp, N. A., Lazic, S. E., Lidster, K., MacCallum, C. J., Macleod, M., … Würbel, H. (2020). The ARRIVE guidelines 2.0: Updated guidelines for reporting animal research. PLoS Biology, 18(7), e3000410. https://doi.org/10.1371/journal.pbio.3000410
Quinton, L., Girard, E., Maiga, A., Rekik, M., Lluel, P., Masuyer, G., Larregola, M., Marquer, C., Ciolek, J., Magnin, T., Wagner, R., Molgó, J., Thai, R., Fruchart-Gaillard, C., Mourier, G., Chamot-Rooke, J., Ménez, A., Palea, S., Servent, D., & Gilles, N. (2010). Isolation and pharmacological characterization of AdTx1, a natural peptide displaying specific insurmountable antagonism of the α1A-adrenoceptor. British Journal of Pharmacology, 159, 316–325. https://doi.org/10.1111/j.1476-5381.2009.00532.x
Rouget, C., Quinton, L., Maïga, A., Gales, C., Masuyer, G., Malosse, C., Chamot-Rooke, J., Thai, R., Mourier, G., de Pauw, E., Gilles, N., & Servent, D. (2010). Identification of a novel snake peptide toxin displaying high affinity and antagonist behaviour for the α2-adrenoceptors. British Journal of Pharmacology, 161, 1361–1374. https://doi.org/10.1111/j.1476-5381.2010.00966.x
Schweitz, H., Heurteaux, C., Bois, P., Moinier, D., Romey, G., & Lazdunski, M. (1994). Calcicludine, a venom peptide of the Kunitz-type protease inhibitor family, is a potent blocker of high-threshold Ca2+ channels with a high affinity for L-type channels in cerebellar granule neurons. Proceedings of the National Academy of Sciences, 91, 878–882. https://doi.org/10.1073/pnas.91.3.878
Shafqat, J., Zaidi, Z. H., & Jörnvall, H. (1990). Purification and characterization of a chymotrypsin Kunitz inhibitor type of polypeptide from the venom of cobra (Naja naja naja). FEBS Letters, 275, 6–8. https://doi.org/10.1016/0014-5793(90)81426-O
Strydom, D. J., & Joubert, F. J. (1981). The amino acid sequence of a weak trypsin inhibitor B from Dendroaspis polylepis polylepis (black mamba) venom. Hoppe-Seyler's Zeitschrift für Physiologische Chemie, 362, 1377–1384. https://doi.org/10.1515/bchm2.1981.362.2.1377
Sun, D., Yu, Y., Xue, X., Pan, M., Wen, M., Li, S., Qu, Q., Li, X., Zhang, L., Li, X., Liu, L., Yang, M., & Tian, C. (2018). Cryo-EM structure of the ASIC1a–mambalgin-1 complex reveals that the peptide toxin mambalgin-1 inhibits acid-sensing ion channels through an unusual allosteric effect. Cell Discovery, 4, 1–11. https://doi.org/10.1038/s41421-018-0026-1
Zhou, X. D., Jin, Y., Lu, Q. M., Li, D. S., Zhu, S. W., Wang, W. Y., & Xiong, Y. L. (2004). Purification, characterization and primary structure of a chymotrypsin inhibitor from Naja atra venom. Comparative Biochemistry and Physiology Part B: Biochemistry and Molecular Biology, 137, 219–224. https://doi.org/10.1016/j.cbpc.2003.11.007