In vivo assembly and large-scale purification of a GPCR - Gα fusion with Gβγ, and characterization of the active complex.

ABSTRACT: G protein coupled receptors (GPCRs) are central players in recognizing a variety of stimuli to mediate diverse cellular responses. This myriad of functions is accomplished by their modular interactions with downstream intracellular transducers, such as heterotrimeric G proteins and arrestins. Assembling a specific GPCR-G protein pair as a purified complex for their structural and functional investigations remains a challenging task, however, because of the low affinity of the interaction. Here, we optimized fusion constructs of the Gα subunit of the heterotrimeric G protein and engineered versions of rat Neurotensin receptor 1 (NTR1), coexpressed and assembled in vivo with Gβ and Gγ. This was achieved by using the baculovirus-based MultiBac system. We thus generated a functional receptor-G

SUBMITTER: Kumar A 

PROVIDER: S-EPMC6324789 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature