Rezaei K, Saar K, Soomets U, Valkna A, Näsman J, Zorko M, Akerman K, Schroeder T, Bartfai T, Langel U
Department of Neurochemistry and Neurotoxicology, Stockholm University, Stockholm, S-10691, Sweden.
Neuropeptides. 2000 Feb;34(1):25-31. doi: 10.1054/npep.1999.0782.
To determine the domains essential for G-protein coupling of the human galanin receptor type 1 (GalR1), we have used both GalR1 mutants and synthetic receptor-derived peptides in(125)I-galanin and [(35)S]-GTPgammaS binding studies. Replacement of potential phosphorylation sites by Leu in the third intracellular loop (IC3) of GalR1 did not affect K(D)values for the receptor. Peptides derived form the IC3 loop, and especially the N-terminal part of it were able to increase the rate of [(35)S]-GTPgammaS binding to the trimeric Gialpha1beta1gamma2, but not to Gsalphabeta1gamma2, whereas the peptides corresponding to the IC1 and IC2 loops had no such effect. IC3 loop peptides also inhibited the binding of(125)I-galanin to GalR1 in membranes from Rin m5F cells. Our results suggest that the IC3 loop of GalR1, especially its N-terminal part, defines the coupling of the receptor to the Gialpha1beta1gamma2 protein and consequently, to the signal transduction cascade.
为了确定人1型甘丙肽受体（GalR1）与G蛋白偶联所必需的结构域，我们在¹²⁵I-甘丙肽和[³⁵S]-GTPγS结合研究中使用了GalR1突变体和合成的受体衍生肽。在GalR1的第三个细胞内环（IC3）中用亮氨酸取代潜在的磷酸化位点并不影响该受体的解离常数（KD）值。源自IC3环的肽，尤其是其N端部分，能够增加[³⁵S]-GTPγS与三聚体Gialpha1beta1gamma2的结合速率，但不能增加与Gsalphabeta1gamma2的结合速率，而对应于IC1和IC2环的肽则没有这种作用。IC3环肽还抑制¹²⁵I-甘丙肽与Rin m5F细胞膜中GalR1的结合。我们的结果表明，GalR1的IC3环，尤其是其N端部分，决定了该受体与Gialpha1beta1gamma2蛋白的偶联，从而决定了与信号转导级联反应的偶联。
