Functional coupling with Gαq and Gαi1 protein subunits promotes high‐affinity agonist binding to the neurotensin receptor NTS‐1 expressed in Escherichia coli

FEBS Letters

Hermans, Emmanuel¹  Grisshammer, Reinhard² 

[1] Laboratory of Pharmacology, Université Catholique de Louvain 54.10, Avenue Hippocrate 54, 1200 Brussels, Belgium;MRC Laboratory of Molecular Biology, Hills Road, Cambridge CB2 2QH, UK

关键词: Neurotensin receptor;    NTS-1;    G-protein α subunit;    Guanosine 5′-O-(3-thiotriphosphate);    Seven-helix G-protein-coupled receptor–Gα fusion;    Escherichia coli;    CHO;    Chinese hamster ovary;    Gi/q;    Gαi1 subunit in which the six C-terminal amino acid residues were replaced with the six C-terminal residues of Gαq;    Gi/s;    Gαi1 subunit containing the six C-terminal amino acid residues of Gαs;    GPCR;    seven-helix G-protein-coupled receptor;    GTPγS;    guanosine 5′-O-(3-thiotriphosphate);    MBP;    Escherichia coli maltose binding protein;    NT;    neurotensin;    NTR;    high-affinity neurotensin receptor NTS-1;    PTX;    pertussis toxin;    TrxA;    Escherichia coli thioredoxin;   

DOI  :  10.1016/S0014-5793(01)02281-5

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

To analyze the coupling of Gα subunits to the rat neurotensin receptor NTS-1 (NTR), fusion proteins were expressed in Escherichia coli with various Gα subunits covalently linked to the receptor C-terminus. The presence of Gα_q or Gα_i/q, in which the six C-terminal residues of Gα_i1 were replaced with those from Gα_q, increased the percentage of receptors in the agonist high-affinity state. This effect was less pronounced for wild-type Gα_i1 and not observed for Gα_i/s. Functional coupling of neurotensin receptor to Gα was demonstrated by neurotensin-induced [³⁵S]GTPγS binding for the Gα_q, Gα_i/q and Gα_i1 subunits, but not for Gα_i/s. Our results extend previous findings of the dual coupling of NTR to pertussis toxin-sensitive and -insensitive G-proteins in Chinese hamster ovary cells with preference for the latter.

【 授权许可】

Unknown