Interactions of endothelin receptor subtypes A and B with Gi, Go, and Gq in reconstituted phospholipid vesicles

被引:49
作者
Doi, T
Sugimoto, H
Arimoto, N
Hiroaki, Y
Fujiyoshi, Y
机构
[1] Matsushita Elect Ind Co Ltd, Int Inst Adv Res, Kyoto 6190237, Japan
[2] Kyoto Univ, Grad Sch Sci, Dept Biophys, Sakyo Ku, Kyoto 6068502, Japan
关键词
D O I
10.1021/bi981919m
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
To understand the biochemical basis for the functional divergence of the human endothelin receptor subtypes A (ETAR) and B (ETBR), they were expressed, purified from insect Sf9 cells, and reconstituted into phospholipid vesicles with the G(o), G(q), and G(i) proteins. For each G protein, a unique pattern of reactivity was observed with the different receptor subtypes. Both ETAR and ETBR activated G(o) to a similar maximal extent, and both subtypes activated G(q) with similar EC50 values; however, the ETAR displayed a 2-3-fold higher maximal extent of activation. In contrast, both subtypes activated G(i) to a similar maximal extent, but the ETAR displayed a 4-fold higher EC50 value as compared to the ETBR. To test whether these coupling specificities are influenced by C-terminal palmitoylation of the receptor, we mutated a cluster of-cysteine residues near the end of the seventh transmembrane helix in both receptors. While the cysteine mutations in the ETBR resulted in a partially palmitoylated receptor, the replacement of these cysteine residues in the ETAR yielded a mostly palmitoyl-deficient receptor and had no effect on G(o) activation, but caused a reduction in the extents of G(i) and G(q) stimulation. Together, these studies provide important insights into the specificity of G protein coupling in the endothelin receptors. The ability to discriminate between the different G proteins under various physiological conditions may be a key element in the selection of distinct signal transduction pathways by the two receptor subtypes.
引用
收藏
页码:3090 / 3099
页数:10
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