Establishment Of Bile Acid Receptor TGR5 Biased Ligand Screening Model And Screening Of TGR5 Agonist

G protein-coupled receptors are currently the most successful family of drug targets.They regulate intracellular signals through G protein signaling pathway and GRK and ?-arrestin signaling pathways,thereby mediating a series of physiological and pathological processes.TGR5 is used as a target for the treatment of diabetes,obesity,inflammation,cancer and other diseases,and provides the impetus for the development of TGR5 agonists and antagonists.The activation of TGR5 can lead to an increase in intracellular cAMP content,which can play a role in reducing blood glucose,anti-inflammatory and promoting energy metabolism.However,the role of TGR5's ?-arrestin signaling pathway in disease regulation needs further study.In this paper,we construct a model for detecting the interaction between TGR5 and ?-arrestin by constructing a fusion expression plasmid of LgBit/SmBit fragment and TGR5 and ?-arrestin1/2 fusion plasmid.First,the cAMP experiment was used to verify the integrity of the TGR5 fusion plasmid,and then TGR5-LgBit/SmBit and ?-arrestin1/2-LgBit/SmBit were combined into HEK293 cells,then use the NanoBit experiment to detect.According to the signal,a model of the interaction between TGR5 and ?-arrestin was established,that is,the combination of TGR5-LgBit and SmBit-?-arrestin1/2.In addition,the ?2AR-LgBit and A2AR-LgBit fusion plasmids were constructed and transferred to HEK293 cells together with SmBit-?-arrestin1/2,and then tested by NanoBit experiment.The detection results were consistent with literature reports.This shows that the model is not only suitable for the detection of the interaction between TGR5 and ?-arrestin,but also for the detection of the interaction of other GPCRs with ?-arrestin.And it shows that the model is accurate and reliable,it provides a new verification method for ligand screening,and can be used in future research on GPCR signal transduction pathways and biased agonists,which lays a certain foundation for further clarification of GPCR signal transduction research.Foundation of methodology.The second part of this paper is to use the cAMP experiment to construct a screening model of TGR5 small molecule agonists.From the structurally modified compounds,we obtained some compounds with better agonistic activity and worthy of further study.This provides a basis for TGR5 agonist to become a drug candidate for diabetes.