title: Biochemical characterization of a heterotrimeric G(i)-protein activator peptide designed from the junction between the intracellular third loop and sixth transmembrane helix in the m4 muscarinic acetylcholine receptor.
abstract: Muscarinic acetylcholine receptors (mAChRs) are G-protein coupled receptors (GPCRs) that are activated by acetylcholine released from parasympathetic nerves. The mAChR family comprises 5 subtypes, m1-m5, each of which has a different coupling selectivity for heterotrimeric GTP-binding proteins (G-proteins). m4 mAChR specifically activates the Gi/o family by enhancing the guanine nucleotide exchange factor (GEF) reaction with the Gα subunit through an interaction that occurs via intracellular segments. Here, we report that the m4 mAChR mimetic peptide m4i3c(14)Gly, comprising 14 residues in the junction between the intracellular third loop (i3c) and transmembrane helix VI (TM-VI) extended with a C-terminal glycine residue, presents GEF activity toward the Gi1 α subunit (Gαi1). The m4i3c(14)Gly forms a stable complex with guanine nucleotide-free Gαi1 via three residues in the VTI(L/F) motif, which is conserved within the m2/4 mAChRs. These results suggest that this m4 mAChR mimetic peptide, which comprises the amino acid of the mAChR intracellular segments, is a useful tool for understanding the interaction between GPCRs and G-proteins.
authors.affiliation: Graduate School of Science and Technology, Gunma University, 1-5-1 Tenjin-cho, Kiryu, Gunma, 376-8515, Japan. Electronic address: terawaki@gunma-u.ac.jp.
authors.forename: Shin-ichi
authors.initials: S
authors.lastname: Terawaki
authors.affiliation: Graduate School of Science and Technology, Gunma University, 1-5-1 Tenjin-cho, Kiryu, Gunma, 376-8515, Japan.
authors.forename: Rina
authors.initials: R
authors.lastname: Matsubayashi
authors.affiliation: Graduate School of Science and Technology, Gunma University, 1-5-1 Tenjin-cho, Kiryu, Gunma, 376-8515, Japan.
authors.forename: Kanako
authors.initials: K
authors.lastname: Hara
authors.affiliation: Graduate School of Science and Technology, Gunma University, 1-5-1 Tenjin-cho, Kiryu, Gunma, 376-8515, Japan.
authors.forename: Tatsuki
authors.initials: T
authors.lastname: Onozuka
authors.affiliation: Department of Biochemistry, Kitasato University School of Medicine, 1-15-1 Kitasato, Minami-ku, Sagamihara, Kanagawa, 252-0374, Japan.
authors.forename: Toshiyuki
authors.initials: T
authors.lastname: Kohno
authors.affiliation: Graduate School of Science and Technology, Gunma University, 1-5-1 Tenjin-cho, Kiryu, Gunma, 376-8515, Japan.
authors.forename: Kaori
authors.initials: K
authors.lastname: Wakamatsu
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D040761:Biomimetic Materials / Q000737:chemistry / Q000378:metabolism; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006153:Guanosine Diphosphate / Q000378:metabolism; D006801:Humans; D051379:Mice; D008958:Models, Molecular; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D017433:Protein Structure, Secondary; D051381:Rats; D043588:Receptor, Muscarinic M4 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
chemical_list: D020662:Guanine Nucleotide Exchange Factors; D043588:Receptor, Muscarinic M4; D011993:Recombinant Fusion Proteins; D006153:Guanosine Diphosphate; D019206:GTP-Binding Protein alpha Subunits, Gi-Go
keywords: GPCR; Gαi1; Mimetic peptide; Muscarinic acetylcholine receptor