Journal of Shandong University (Health Sciences) ›› 2022, Vol. 60 ›› Issue (9): 85-90.doi: 10.6040/j.issn.1671-7554.0.2022.0368

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Sensitivity of GPCR-G protein fusion sensor to detect the interaction between GPCR and G protein

MA Chuanshun1,2, LIN Hui2, SUN Jinpeng1,2, ZHANG Daolai1   

  1. 1. School of Pharmacy, Binzhou Medical University, Yantai 264003, Shandong, China;
    2. Department of Basic Medical Sciences, Shandong University, Jinan 250012, Shandong, China
  • Published:2022-09-02

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Abstract: Objective GPR56-G protein fusion sensor was used as a prototype to establish a highly sensitive method to detect the interaction between GPCR and G protein. Methods The N-terminus of the G protein α subunit with NanoLuc luciferase(Nluc)was fused to the C-terminus of the GPCR. The fusion protein of both GPCR and G protein α subunit was constructed as the experimental group, and the receptor and G protein were co-expressed as a control group. The constitutive activity of GPR56-Gα12 subunit fusion protein, GPR56 and Gα12 subunit co-expression were compared with bioluminescence resonance energy transfer(BRET)at the same level of receptor expression. The effect of agonist(P19)on the interaction of GPR56-Gα12 subunit fusion protein was detected with BRET. Results The BRET ratio results showed that the GPR56-Gα12 subunit fusion protein had stronger constitutive activity than the control group co-expressed with GPR56 and Gα12 subunits(F=424.7, P<0.001). Compared with the control group, the experimental group had decreased half effective concentration(EC50)of P19 activation (t=13.36, P<0.001). The GPR56-Gα12 subunit fusion protein was more sensitive to P19 and had a stronger affinity. Conclusion The GPCR-Gα subunit fusion protein system is a sensitive method to detect the interaction between GPCRs and G protein.

Key words: GPCR-Gα subunit fusion protein, GPR56, Gα12, Constitutive activity, P19

CLC Number: 

  • Q5-33
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