title: Conversed mutagenesis of an inactive peptide to ASIC3 inhibitor for active sites determination.
abstract: Peptide Ugr9-1 from the venom of sea anemone Urticina grebelnyi selectively inhibits the ASIC3 channel and significantly reverses inflammatory and acid-induced pain in vivo. A close homolog peptide Ugr 9-2 does not have these features. To find the pharmacophore residues and explore structure-activity relationships of Ugr 9-1, we performed site-directed mutagenesis of Ugr 9-2 and replaced several positions by the corresponding residues from Ugr 9-1. Mutant peptides Ugr 9-2 T9F and Ugr 9-2 Y12H were able to inhibit currents of the ASIC3 channels 2.2 times and 1.3 times weaker than Ugr 9-1, respectively. Detailed analysis of the spatial models of Ugr 9-1, Ugr 9-2 and both mutant peptides revealed the presence of the basic-aromatic clusters on opposite sides of the molecule, each of which is responsible for the activity. Additionally, Ugr9-1 mutant with truncated N- and C-termini retained similar with the Ugr9-1 action in vitro and was equally potent in vivo model of thermal hypersensitivity. All together, these results are important for studying the structure-activity relationships of ligand-receptor interaction and for the future development of peptide drugs from animal toxins.
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, ul. Miklukho-Maklaya 16/10, 117997 Moscow, Russia. Electronic address: osmadim@gmail.com.
authors.forename: Dmitry I
authors.initials: DI
authors.lastname: Osmakov
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, ul. Miklukho-Maklaya 16/10, 117997 Moscow, Russia.
authors.forename: Sergey G
authors.initials: SG
authors.lastname: Koshelev
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, ul. Miklukho-Maklaya 16/10, 117997 Moscow, Russia.
authors.forename: Yaroslav A
authors.initials: YA
authors.lastname: Andreev
authors.affiliation: Branch of the Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 6 Nauki Avenue, Pushchino, 142290 Moscow Region, Russia.
authors.forename: Igor A
authors.initials: IA
authors.lastname: Dyachenko
authors.affiliation: Branch of the Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 6 Nauki Avenue, Pushchino, 142290 Moscow Region, Russia.
authors.forename: Dmitry A
authors.initials: DA
authors.lastname: Bondarenko
authors.affiliation: Branch of the Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 6 Nauki Avenue, Pushchino, 142290 Moscow Region, Russia.
authors.forename: Arkadii N
authors.initials: AN
authors.lastname: Murashev
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, ul. Miklukho-Maklaya 16/10, 117997 Moscow, Russia.
authors.forename: Eugene V
authors.initials: EV
authors.lastname: Grishin
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, ul. Miklukho-Maklaya 16/10, 117997 Moscow, Russia.
authors.forename: Sergey A
authors.initials: SA
authors.lastname: Kozlov
mesh_terms: D062646:Acid Sensing Ion Channel Blockers / Q000737:chemistry*; D062565:Acid Sensing Ion Channels / Q000737:chemistry*; D000818:Animals; D020134:Catalytic Domain; D003064:Cnidarian Venoms / Q000737:chemistry*; D006801:Humans; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D017434:Protein Structure, Tertiary; D020539:Sequence Analysis, Protein; D013329:Structure-Activity Relationship; D014982:Xenopus laevis
chemical_list: D062646:Acid Sensing Ion Channel Blockers; D062565:Acid Sensing Ion Channels; D003064:Cnidarian Venoms
keywords: ASIC; Inflammation; Mutagenesis; Peptides; Sea anemone venom