Method for measuring the modulation of the activation of a G protein-coupled receptor with GTP analogues

A method for determining the capacity of a molecule for modulating the activation of a G protein-coupled receptor (GPCR), said method comprising the following steps:

• a) introducing, in a first container:
• a membrane preparation bearing one or more GPCRs and one or more alpha G proteins,
• a source of nonhydrolyzable or slowly hydrolyzable GTP labeled with a first member of a pair of resonance energy transfer (RET) partners,
• a ligand of the alpha subunit of a G protein (alpha G-protein) labeled with a second member of the pair of RET partners, said ligand being capable of binding to the full alpha G-protein bound to the nonhydrolyzable or slowly hydrolyzable GTP labeled with the first member of a pair of RET partners,
• optionally a GPCR agonist;
• b) measuring the RET signal emitted in the first container;
• c) introducing (i) in a second container, the same reagents as in step a) and the molecule to be assayed or (ii) in the first container, the molecule to be assayed;
• d) measuring the RET signal emitted in the second container or in the first container obtained in step c);
• e) comparing the signals obtained in steps b) and d), a modulation of the signal obtained in step d) relative to that obtained in step b) indicates that the molecule to be tested is capable of modulating the activation of the GPCR, wherein the ligand of the alpha G-protein is an antibody or an antibody fragment capable of binding to the alpha G-protein, which comprises:
• a variable domain of a heavy chain comprising a CDR1 of amino acid sequence SEQ ID NO:
• 2, a CDR2 of amino acid sequence SEQ ID NO:
• 3, and a CDR3 of amino acid sequence SEQ ID NO:
• 4, and
• a variable domain of a light chain comprising a CDR1 of amino acid sequence SEQ ID NO:
• 5, a CDR2 of amino acid sequence DTS, and a CDR3 of amino acid sequence SEQ ID NO:
• 6.