title: Structural Insight into Specificity of Interactions between Nonconventional Three-finger Weak Toxin from Naja kaouthia (WTX) and Muscarinic Acetylcholine Receptors.
abstract: Weak toxin from Naja kaouthia (WTX) belongs to the group of nonconventional "three-finger" snake neurotoxins. It irreversibly inhibits nicotinic acetylcholine receptors and allosterically interacts with muscarinic acetylcholine receptors (mAChRs). Using site-directed mutagenesis, NMR spectroscopy, and computer modeling, we investigated the recombinant mutant WTX analogue (rWTX) which, compared with the native toxin, has an additional N-terminal methionine residue. In comparison with the wild-type toxin, rWTX demonstrated an altered pharmacological profile, decreased binding of orthosteric antagonist N-methylscopolamine to human M1- and M2-mAChRs, and increased antagonist binding to M3-mAChR. Positively charged arginine residues located in the flexible loop II were found to be crucial for rWTX interactions with all types of mAChR. Computer modeling suggested that the rWTX loop II protrudes to the M1-mAChR allosteric ligand-binding site blocking the entrance to the orthosteric site. In contrast, toxin interacts with M3-mAChR by loop II without penetration into the allosteric site. Data obtained provide new structural insight into the target-specific allosteric regulation of mAChRs by "three-finger" snake neurotoxins.
authors.affiliation: From the Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 16/10 Miklukho-Maklaya Street, 117997 Moscow, Russia, the Lomonosov Moscow State University, 119991 Moscow, Russia, ekaterina-lyukmanova@yandex.ru.
authors.forename: Ekaterina N
authors.initials: EN
authors.lastname: Lyukmanova
authors.affiliation: From the Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 16/10 Miklukho-Maklaya Street, 117997 Moscow, Russia, the Lomonosov Moscow State University, 119991 Moscow, Russia, the Moscow Institute of Physics and Technology, Institutskiy Pereulok 9, Dolgoprudny, Moscow Region 141700, Russian.
authors.forename: Zakhar O
authors.initials: ZO
authors.lastname: Shenkarev
authors.affiliation: From the Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 16/10 Miklukho-Maklaya Street, 117997 Moscow, Russia, the Lomonosov Moscow State University, 119991 Moscow, Russia.
authors.forename: Mikhail A
authors.initials: MA
authors.lastname: Shulepko
authors.affiliation: From the Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 16/10 Miklukho-Maklaya Street, 117997 Moscow, Russia, the Lomonosov Moscow State University, 119991 Moscow, Russia.
authors.forename: Alexander S
authors.initials: AS
authors.lastname: Paramonov
authors.affiliation: From the Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 16/10 Miklukho-Maklaya Street, 117997 Moscow, Russia, the Lomonosov Moscow State University, 119991 Moscow, Russia.
authors.forename: Anton O
authors.initials: AO
authors.lastname: Chugunov
authors.affiliation: the Institute of Physiology, Academy of Sciences of the Czech Republic (Public Research Institution), 14220 Prague, Czech Republic, and.
authors.forename: Helena
authors.initials: H
authors.lastname: Janickova
authors.affiliation: the Institute of Physiology, Academy of Sciences of the Czech Republic (Public Research Institution), 14220 Prague, Czech Republic, and.
authors.forename: Eva
authors.initials: E
authors.lastname: Dolejsi
authors.affiliation: the Institute of Physiology, Academy of Sciences of the Czech Republic (Public Research Institution), 14220 Prague, Czech Republic, and.
authors.forename: Vladimir
authors.initials: V
authors.lastname: Dolezal
authors.affiliation: From the Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 16/10 Miklukho-Maklaya Street, 117997 Moscow, Russia.
authors.forename: Yuri N
authors.initials: YN
authors.lastname: Utkin
authors.affiliation: From the Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 16/10 Miklukho-Maklaya Street, 117997 Moscow, Russia.
authors.forename: Victor I
authors.initials: VI
authors.lastname: Tsetlin
authors.affiliation: From the Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 16/10 Miklukho-Maklaya Street, 117997 Moscow, Russia, the Moscow Institute of Physics and Technology, Institutskiy Pereulok 9, Dolgoprudny, Moscow Region 141700, Russian.
authors.forename: Alexander S
authors.initials: AS
authors.lastname: Arseniev
authors.affiliation: From the Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 16/10 Miklukho-Maklaya Street, 117997 Moscow, Russia, the Moscow Institute of Physics and Technology, Institutskiy Pereulok 9, Dolgoprudny, Moscow Region 141700, Russian.
authors.forename: Roman G
authors.initials: RG
authors.lastname: Efremov
authors.affiliation: From the Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 16/10 Miklukho-Maklaya Street, 117997 Moscow, Russia, the Lomonosov Moscow State University, 119991 Moscow, Russia.
authors.forename: Dmitry A
authors.initials: DA
authors.lastname: Dolgikh
authors.affiliation: From the Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 16/10 Miklukho-Maklaya Street, 117997 Moscow, Russia, the Lomonosov Moscow State University, 119991 Moscow, Russia.
authors.forename: Mikhail P
authors.initials: MP
authors.lastname: Kirpichnikov
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D004546:Elapid Venoms / Q000737:chemistry*; D017815:Elapidae; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D009498:Neurotoxins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019906:Nuclear Magnetic Resonance, Biomolecular; D011485:Protein Binding; D011487:Protein Conformation; D011976:Receptors, Muscarinic / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid
chemical_list: D004546:Elapid Venoms; D009498:Neurotoxins; D011976:Receptors, Muscarinic
keywords: G protein-coupled receptor (GPCR); computer modeling; nuclear magnetic resonance (NMR); protein dynamic; recombinant protein expression; site-directed mutagenesis; snake neurotoxin