Shibata T, Suzuki C, Ohnishi J, Murakami K, Miyazaki H
Institute of Applied Biochemistry, University of Tsukuba, Ibaraki, Japan.
Biochem Biophys Res Commun. 1996 Jan 5;218(1):383-9. doi: 10.1006/bbrc.1996.0067.
Previous mutagenesis studies of angiotensin II (Ang II) receptor type 1 (AT1) have focused on determining the regions responsible for Gq coupling using the rat AT1 receptor. We created human AT1 receptor mutants, expressed them in COS-7 cells, and identified the domains crucial for Gi coupling as well as for Gq coupling. Substitution of Asp125, Arg126, Tyr127, and Met134 by Gly, Gly, Ala, and Ala in the highly conserved sequence of the second intracellular loop in most G protein-coupled receptors provided a mutant AT1 receptor which lost the ability to couple to both Gq and Gi with no impairment in its binding to Ang II. A truncated mutant lacking the carboxyl terminal 50 residues was completely deficient in coupling to Gi, whereas it retained full ability to bind to Gq, in contrast to the rat AT1 receptor. These findings demonstrate that the cytoplasmic tail in the human AT1 receptor is the determinant of specific Gi coupling.
先前对1型血管紧张素II（Ang II）受体（AT1）的诱变研究集中于使用大鼠AT1受体来确定负责与Gq偶联的区域。我们构建了人AT1受体突变体，在COS-7细胞中表达它们，并确定了对Gi偶联以及对Gq偶联至关重要的结构域。在大多数G蛋白偶联受体的第二个细胞内环的高度保守序列中，将Asp125、Arg126、Tyr127和Met134替换为Gly、Gly、Ala和Ala，得到了一个突变型AT1受体，该受体失去了与Gq和Gi偶联的能力，但其与Ang II的结合未受损害。与大鼠AT1受体相反，一个缺少羧基末端50个残基的截短突变体完全缺乏与Gi偶联的能力，而它保留了与Gq结合的全部能力。这些发现表明，人AT1受体中的细胞质尾巴是特异性Gi偶联的决定因素。
