Mutational and computational analysis of the alpha(1b)-adrenergic receptor. Involvement of basic and hydrophobic residues in receptor activation and G protein coupling.

Details

Serval ID
serval:BIB_8CEB5BDDCCBA
Type
Article: article from journal or magazin.
Collection
Publications
Institution
Title
Mutational and computational analysis of the alpha(1b)-adrenergic receptor. Involvement of basic and hydrophobic residues in receptor activation and G protein coupling.
Journal
Journal of Biological Chemistry
Author(s)
Greasley P.J., Fanelli F., Scheer A., Abuin L., Nenniger-Tosato M., DeBenedetti P.G., Cotecchia S.
ISSN
0021-9258 (Print)
ISSN-L
0021-9258
Publication state
Published
Issued date
2001
Peer-reviewed
Oui
Volume
276
Number
49
Pages
46485-46494
Language
english
Abstract
To investigate their role in receptor coupling to G(q), we mutated all basic amino acids and some conserved hydrophobic residues of the cytosolic surface of the alpha(1b)-adrenergic receptor (AR). The wild type and mutated receptors were expressed in COS-7 cells and characterized for their ligand binding properties and ability to increase inositol phosphate accumulation. The experimental results have been interpreted in the context of both an ab initio model of the alpha(1b)-AR and of a new homology model built on the recently solved crystal structure of rhodopsin. Among the twenty-three basic amino acids mutated only mutations of three, Arg(254) and Lys(258) in the third intracellular loop and Lys(291) at the cytosolic extension of helix 6, markedly impaired the receptor-mediated inositol phosphate production. Additionally, mutations of two conserved hydrophobic residues, Val(147) and Leu(151) in the second intracellular loop had significant effects on receptor function. The functional analysis of the receptor mutants in conjunction with the predictions of molecular modeling supports the hypothesis that Arg(254), Lys(258), as well as Leu(151) are directly involved in receptor-G protein interaction and/or receptor-mediated activation of the G protein. In contrast, the residues belonging to the cytosolic extensions of helices 3 and 6 play a predominant role in the activation process of the alpha(1b)-AR. These findings contribute to the delineation of the molecular determinants of the alpha(1b)-AR/G(q) interface.
Keywords
Amino Acid Sequence, Animals, COS Cells, Cricetinae, GTP-Binding Proteins/metabolism, Models, Molecular, Molecular Sequence Data, Mutagenesis, Polymerase Chain Reaction, Protein Conformation, Receptors, Adrenergic, alpha-1/chemistry, Receptors, Adrenergic, alpha-1/genetics
Pubmed
Web of science
Open Access
Yes
Create date
24/01/2008 11:05
Last modification date
20/08/2019 14:51
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