Mutation of a single TMVI residue, Phe(282), in the beta(2)-adrenergic receptor results in structurally distinct activated receptor conformations

Songhai Chen; Fang Lin; Ming Xu; R Peter Riek; Jiri Novotny; Robert M Graham

doi:10.1021/bi012189c

Back

Mutation of a single TMVI residue, Phe(282), in the beta(2)-adrenergic receptor results in structurally distinct activated receptor conformations

Journal article

Peer reviewed

Mutation of a single TMVI residue, Phe(282), in the beta(2)-adrenergic receptor results in structurally distinct activated receptor conformations

Songhai Chen, Fang Lin, Ming Xu, R Peter Riek, Jiri Novotny and Robert M Graham

Biochemistry (Easton), Vol.41(19), pp.6045-6053

05/14/2002

DOI: 10.1021/bi012189c

PMID: 11993999

View Online

Abstract

We showed previously that Phe(303) in transmembrane segment (TM) VI of the alpha(1B)-adrenergic receptor (alpha(1B)-AR), a residue conserved in many G protein-coupled receptors (GPCRs), is critically involved in coupling agonist binding with TM helical movement and G protein activation. Here the equivalent residue, Phe(282), in the beta(2)-AR was evaluated by mutation to glycine, asparagine, alanine, or leucine. Except for F282N, which exhibits attenuated basal and maximal isoproterenol stimulation, the Phe(282) mutants display varying degrees of constitutive activity (F282L > F282A > F282G), and as shown by the results of substituted cysteine accessibility method (SCAM) studies, induce movement of endogenous cysteine(s) into the water-accessible ligand-binding pocket. For F282A, movement is confined to Cys(285) in TMVI, whereas F282L induces movement of both Cys(285) in TMVI and Cys(327) in TMVII. Further, engineered cysteine-sensor studies indicate that F282L causes movement of TMVI, both above and below an apparent kink-inducing TMVI proline (Pro(288)), whereas that due to F282A is confined to the domain below Pro(288). A plausible interpretation of these data is that receptor activation involves rigid body movement of TMVI which, because of its Pro(288)-induced kink, acts as a pivot to transduce and amplify the agonist-induced conformational change in the upper domain, to a change in the lower domain required for productive receptor-G protein coupling.

Molecular Sequence Data

Receptors, Adrenergic, beta-2 - chemistry

Phenylalanine - chemistry

Computer Simulation

Conserved Sequence

Recombinant Proteins - metabolism

Amino Acid Sequence

Cricetinae

Mutagenesis, Site-Directed

Protein Structure, Secondary

Receptors, Adrenergic, beta-2 - genetics

Models, Molecular

Recombinant Proteins - chemistry

Recombinant Proteins - genetics

Binding Sites - genetics

Receptors, Adrenergic, beta-2 - metabolism

Sequence Homology, Amino Acid

Animals

Ligands

Protein Conformation

Kinetics

In Vitro Techniques

COS Cells

Amino Acid Substitution

GTP-Binding Proteins - metabolism

Details

Title: Subtitle: Mutation of a single TMVI residue, Phe(282), in the beta(2)-adrenergic receptor results in structurally distinct activated receptor conformations
Creators: Songhai Chen - Molecular Cardiology and Biocomputing Units, Victor Chang Cardiac Research Institute, St. Vincent's Hospital, Darlinghurst, NSW 2010, Australia. songhai.chen@mcmail.vanderbilt.edu
Fang Lin
Ming Xu
R Peter Riek
Jiri Novotny
Robert M Graham
Resource Type: Journal article
Publication Details: Biochemistry (Easton), Vol.41(19), pp.6045-6053
DOI: 10.1021/bi012189c
PMID: 11993999
NLM abbreviation: Biochemistry
ISSN: 0006-2960
eISSN: 1520-4995
Publisher: United States
Language: English
Date published: 05/14/2002
Academic Unit: Anatomy and Cell Biology; Iowa Neuroscience Institute; Craniofacial Anomalies Research Center; Fraternal Order of Eagles Diabetes Research Center; Neuroscience and Pharmacology; Internal Medicine
Record Identifier: 9984040446802771

Metrics

17 Record Views

29 Times Cited - Web of Science