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Genetic interactions among the transmembrane segments of the G protein coupled receptor encoded by the yeast STE2 gene.

Abstract：

:G protein coupled receptors (GPCRs) are integral membrane proteins that mediate cellular responses to a wide variety of extracellular signals. However, the structural basis for activation of this class of receptors by ligand binding is not well understood. We report here the use of a systematic genetic protocol for identifying interactions among the seven transmembrane helices of the GPCR responsible for cellular responses to the alpha-mating pheromone of the yeast Saccharomyces cerevisiae. Random mutations were introduced into the region of the STE2 gene encoding the third transmembrane segment of the alpha-factor receptor, followed by screening for loss of signaling. The limited spectrum of non-conservative mutations recovered, including removal of the only negatively charged side-chain in the transmembrane region, indicates that most substitutions in the third transmembrane segment do not affect receptor function. Three second-site intragenic suppressors of these initial mutations were isolated following mutagenesis of the remaining six transmembrane segments. One of these suppressors, Y266C in the sixth transmembrane segment, is allele specific and shows non-additivity of phenotypes indicative of a physical interaction between the third and sixth transmembrane regions of the receptor. A second suppressor, M218T in the fifth transmembrane segment, exhibits only partial allele specificity. A third suppressor, R58G, in the first transmembrane segment, suppresses a variety of starting alleles and appears to cause global stabilization of the receptor. Analysis of these suppressors and additional alleles can provide a database for modeling GPCR structure.

journal_name

J Mol Biol

journal_title

Journal of molecular biology

authors

Sommers CM,Dumont ME

doi

10.1006/jmbi.1996.0816

subject

Has Abstract

pub_date

1997-02-28 00:00:00

pages

559-75

issue

3

eissn

0022-2836

issn

1089-8638

pii

S0022-2836(96)90816-5

journal_volume

266

pub_type

杂志文章

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