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Functional analysis of a human A 1 adenosine receptor/green fluorescent protein/G i1 α fusion protein following stable expression in CHO cells
Author(s) -
Bevan Nicola,
Palmer Tim,
Drmota Tomas,
Wise Alan,
Coote Jim,
Milligan Graeme,
Rees Stephen
Publication year - 1999
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(99)01467-2
Subject(s) - g protein , adenylyl cyclase , pertussis toxin , fusion protein , microbiology and biotechnology , gi alpha subunit , gs alpha subunit , green fluorescent protein , biology , g alpha subunit , protein kinase a , chemistry , biochemistry , receptor , protein subunit , phosphorylation , recombinant dna , gene
Fusion proteins between the human A 1 adenosine receptor and the pertussis toxin resistant (Cys351Gly) mutant of the G‐protein α subunit G i1 α (A1/Gi), and between the human A 1 adenosine receptor, the Aequorea victoria green fluorescent protein (GFP) and Cys351Gly G i1 α (A1/GFP/Gi), were expressed in CHO cells. The agonist NECA caused a stimulation of [ 35 S]GTPγS binding at both fusion proteins with similar concentration dependence as at the native receptor. However in the presence of pertussis toxin NECA stimulation of [ 35 S]GTPγS binding was only seen at the A1/GFP/Gi fusion protein. The regulation of the adenylyl cyclase and MAP kinase effector systems by both fusion proteins was attenuated following pertussis toxin treatment. These studies demonstrate for the first time the characterisation of a fusion protein between a G‐protein coupled receptor, GFP and a G‐protein α subunit.