We found a match
Your institution may have rights to this item. Sign in to continue.
- Title
Interaction of the muscarinic acetylcholine receptor M2 subtype with G protein Gαi/o isotypes and Gβγ subunits as studied with the maltose-binding protein-M2-Gαi/o fusion proteins expressed in Escherichia coli.
- Authors
Ichiyama, Susumu; Nemoto, Reiko; Tanabe, Hiroaki; Haga, Tatsuya
- Abstract
We expressed the fusion proteins of the muscarinic acetylcholine receptor M2 subtype (M2 receptor) with a maltose-binding protein (MBP) and various G protein α subunits (Gαi1–i3/o) at its N- and C-terminals, respectively (MBP-M2-Gαi/o), in Escherichia coli, and examined the effect of G protein βγ subunits (Gβγ) on the receptor–Gα interaction as assessed by agonist- and GDP-dependent [35S]GTPγS binding of the fusion proteins. We found that (i) Gβγ promoted both the agonist-dependent and -independent [35S]GTPγS binding with little effect on the guanine nucleotide-sensitive high-affinity agonist binding, (ii) the specific [35S]GTPγS binding activity was much greater for MBP-M2-GαoA than for MBP-M2-Gαi1–i3 in the absence of Gβγ, whereas Gβγ preferentially promoted the agonist-dependent decrease in the affinity for GDP of MBP-M2-Gαi1–i3 rather than of MBP-M2-GαoA, and (iii) the proportion of agonist-dependent [35S]GTPγS binding was roughly 50% irrespective of species of Gα and the presence or absence of Gβγ. These results demonstrate that receptor-Gα fusion proteins expressed in E. coli could be useful for studies of receptor–G interaction.
- Subjects
MUSCARINIC acetylcholine receptors; G proteins; CHIMERIC proteins; ESCHERICHIA coli; MALTOSE-binding proteins
- Publication
Journal of Biochemistry, 2014, Vol 156, Issue 5, p259
- ISSN
0021-924X
- Publication type
Article
- DOI
10.1093/jb/mvu036