Functional segregation of the highly conserved basic motifs within the third endoloop of the human secretin receptor
Functional segregation of the highly conserved basic motifs within the third endoloop of the human secretin receptor
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DOI：
10.1210/en.142.9.3926
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发表时间：
2001-09-01
期刊：
影响因子：
4.8
通讯作者：
Chow, BKC
中科院分区：
文献类型：
--
作者：
Chan, KYY;Pang, RTK;Chow, BKC
在这项研究中，诱变为基础的战略，以评估的作用，两个高度保守的基序（KLR和RLAR）内的第三内环的人分泌素受体。阻断KLRT和突变Lys 323（（KI）-I-323）显著降低cAMP积累，并且这些突变不影响配体相互作用和细胞表面表达的受体数量。因此，在第三个内环的N末端的KLRT区域，特别是Lys 323，对于G蛋白偶联是重要的。对于RLAR基序，发现在339和342位从Arg取代为Ala（R-339，（342）A）、Glu（R-339，R-E-342）或Ile（（RI）-I-339，342）以及RLAR基序的阻断缺失的受体在促分泌素结合和cAMP产生方面都有缺陷。有趣的是，在Arg 339或Arg 342的相应位置处的单个突变在所有功能测定中响应为野生型人促胰液素受体，这表明在RLAR基序内的任一位置处存在一个Arg足以用于正常的受体功能。这些突变体受体的免疫荧光染色表明，这些精氨酸残基负责表面呈递和/或受体稳定性。
In this study, a mutagenesis-based strategy was employed to assess the roles of two highly conserved motifs (KLR and RLAR) within the third endoloop of the human secretin receptor. Block deletion of KLRT and mutation of Lys323 ((KI)-I-323) significantly reduced cAMP accumulation, and these mutations did not affect ligand interaction and receptor number expressed on the cell surface. Thus, the KLRT region at the N terminus of the third endoloop, particularly Lys323, is important for G protein coupling. For the RLAR motif, receptors with substitutions at positions 339 and 342 from Arg to Ala (R-339, (342)A), Glu (R-339,R- E-342), or Ile ((RI)-I-339, 342) as well as block deletion of the RLAR motif were all found to be defective in both secretin-binding and cAMP production. Interestingly, a single mutation at the corresponding positions of Arg339 or Arg342 responded as the wild-type human secretin receptor in all functional assays, indicating that the presence of one Arg at either position within the RLAR motif is sufficient for a normal receptor function. Immunofluorescent, staining of these mutant receptors showed that these Arg residues are responsible for surface presentation and/or receptor stability.