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Preparation of functional human lysophosphatidic acid receptor 2 using a P9 ∗ expression system and an amphipathic polymer and investigation of its in vitro binding preference to G α proteins

2017; Elsevier BV; Volume: 487; Issue: 1 Linguagem: Inglês

10.1016/j.bbrc.2017.04.025

1090-2104

Seong‐Gu Han, Seung-Il Baek, Tae Jin Son, Hyeongjin Lee, Nam Hyuk Kim, Yeon Gyu Yu,

Cellular transport and secretion

Human lysophosphatidic acid receptor 2 (LPA 2 ), a member of the G-protein coupled receptor family, mediates lysophosphatidic acid (LPA)-dependent signaling by recruiting various G proteins . Particularly, it is directly implicated in the progression of colorectal and ovarian cancer through G protein signaling cascades. To investigate the biochemical binding properties of LPA 2 against various alpha subunits of G protein (G α ), a functional recombinant LPA 2 was overexpressed in E. coli membrane with a P9 ∗ expression system , and the purified protein was stabilized with an amphipathic polymer that had been synthesized by coupling octylamine , glucosamine , and diethyl aminoproylamine at the carboxylic groups of poly-γ-glutamic acid. The purified LPA 2 stabilized with the amphipathic polymer showed selective binding activity to the various G α proteins as well as agonist-dependent dissociation from G αi3 . Understanding the binding properties of LPA 2 against various G α proteins advances the understanding of downstream signaling cascades of LPA 2 . The functional LPA 2 prepared using a P9 ∗ expression system and an amphipathic polymer could also facilitate the development of LPA 2 -targeting drugs.