Document Detail


Hydrophobic amino acid in the i2 loop plays a key role in receptor-G protein coupling.
MedLine Citation:
PMID:  8226735     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Signal transduction of the heptahelical G protein-coupled receptors (GPCRs) involves multiple receptor domains, but a universal consensus domain for coupling has not yet been defined. Alanine mutagenesis scanning was performed on the intracellular loops and the COOH tail of the human muscarinic cholinergic receptor (Hm1) to identify coupling domains. Stimulation of phosphatidylinositol (PI) turnover was determined after transfection of the alanine mutants into U293 human embryonic kidney cells. Alanine substitutions in four regions (loops i1, i2, and NH2 and COOH junctions of i3) impaired coupling efficiency by approximately 50% or more, but the strongest reduction (> 80%) resulted from alanine replacement of a single amino acid, leucine 131. This residue is located in the middle of the second intracellular loop (i2), within the highly conserved GPCR motif (DRYXXV(I)XXPL). The position equivalent to Leu-131 in Hm1 contains a bulky hydrophobic amino acid (L, I, V, M, or F) in nearly all cloned GPCRs. Substitution of Leu-131 with polar amino acids (aspartate and asparagine) also resulted in strongly defective coupling, whereas phenylalanine (found in the equivalent position in the beta 2 adrenoceptor) can replace leucine without losing PI coupling ability of Hm1. Alanine substitution of the corresponding amino acid in the Hm3 receptor (L174A) also inhibited agonist-stimulated PI turnover, while replacing Phe-139 with alanine in the beta 2 adrenoceptor suppressed stimulation of adenylyl cyclase. We propose that a bulky hydrophobic amino acid in the middle of the i2 loop serves as a general site relevant to G protein coupling, whereas coupling selectivity is governed by other receptor domains.
Authors:
O Moro; J Lameh; P Högger; W Sadée
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Publication Detail:
Type:  Comparative Study; Journal Article; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  The Journal of biological chemistry     Volume:  268     ISSN:  0021-9258     ISO Abbreviation:  J. Biol. Chem.     Publication Date:  1993 Oct 
Date Detail:
Created Date:  1993-12-01     Completed Date:  1993-12-01     Revised Date:  2007-11-15    
Medline Journal Info:
Nlm Unique ID:  2985121R     Medline TA:  J Biol Chem     Country:  UNITED STATES    
Other Details:
Languages:  eng     Pagination:  22273-6     Citation Subset:  IM    
Affiliation:
Department of Pharmacy, University of California, San Francisco 94143-0446.
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MeSH Terms
Descriptor/Qualifier:
Amino Acid Sequence
Binding Sites
Carbachol / pharmacology
Cell Line
Consensus Sequence
Cyclic AMP / metabolism
Embryo, Mammalian
GTP-Binding Proteins / chemistry*,  metabolism*
Humans
Isoproterenol / pharmacology
Kidney
Kinetics
Leucine
Molecular Sequence Data
Mutagenesis, Site-Directed
Phosphatidylinositols / metabolism
Point Mutation
Protein Structure, Secondary*
Receptors, Cell Surface / chemistry*,  metabolism
Receptors, Muscarinic / biosynthesis,  chemistry*,  metabolism*
Recombinant Proteins / biosynthesis,  chemistry,  metabolism
Sequence Homology, Amino Acid
Transfection
Grant Support
ID/Acronym/Agency:
GM43102/GM/NIGMS NIH HHS; MH00996/MH/NIMH NIH HHS
Chemical
Reg. No./Substance:
0/Phosphatidylinositols; 0/Receptors, Cell Surface; 0/Receptors, Muscarinic; 0/Recombinant Proteins; 51-83-2/Carbachol; 60-92-4/Cyclic AMP; 61-90-5/Leucine; 7683-59-2/Isoproterenol; EC 3.6.1.-/GTP-Binding Proteins

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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