Structural motifs of importance for the constitutive activity of the orphan 7TM receptor EBI2: Analysis of receptor activation in the absence of an agonist
Structural motifs of importance for the constitutive activity of the orphan 7TM receptor EBI2: Analysis of receptor activation in the absence of an agonist
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DOI：
10.1124/mol.108.049676
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发表时间：
2008-10-01
影响因子：
3.6
通讯作者：
Rosenkilde, Mette M.
中科院分区：
文献类型：
--
作者：
Benned-Jensen, Tau;Rosenkilde, Mette M.
Epstein-Barr诱导受体2 （EBI2）是一种淋巴细胞表达的孤儿7跨膜（7TM）受体，通过G α (i)组成信号，如图所示，例如鸟苷5 '- o -（3-硫代）三磷酸结合。通过对EBI2中29个残基的系统突变分析，确定了两个对构成活性重要的区域。cAMP反应元件结合蛋白转录因子被用来衡量受体活性，并与受体表面表达相关。在TM 6中保守的CW/FxP基序中，PheVI:13 （Phe(257)）和邻近的CysVI:12 （Cys(256)）作为负调控因子，在这些位置上的Ala取代分别使EBI2野生型（wt）的组成活性提高了5.7倍和2.3倍。相比之下，TM-2中的ArgII:20 （Arg(87)）作为正调节因子，替代了Ala，而不是Lys，与wt EBI2相比，其组成活性降低了7倍以上。IleIII:03 （Ile106）距离ArgII: 20仅4埃，在III:03中引入Glu与ArgII: 20产生良好的静电相互作用，活性提高到EBI2的4.4倍。值得注意的是，通过在II:20和III:03中引入Glu和Arg交换这些电荷，与wt EBI2相比，增加了2.7倍，从而挽救了两个信号缺陷单突变，其组成活性降低了3.8- 4.5倍。从药物开发的角度来看，发现这些EBI2激活的分子机制是很重要的，因为它可能有助于合理设计和开发针对EBI2的小分子逆激动剂，这些激动剂被认为是抗病毒或免疫调节疗法的重要作用。
The Epstein-Barr induced receptor 2 (EBI2) is a lymphocyte-expressed orphan seven transmembrane-spanning (7TM) receptor that signals constitutively through G alpha(i), as shown, for instance by guanosine 5 '-O-(3-thio)triphosphate incorporation. Two regions of importance for the constitutive activity were identified by a systematic mutational analysis of 29 residues in EBI2. The cAMP response element-binding protein transcription factor was used as a measure of receptor activity and was correlated to the receptor surface expression. PheVI:13 (Phe(257)), and the neighboring CysVI:12 (Cys(256)), in the conserved CW/FxP motif in TM 6, acted as negative regulators as Ala substitutions at these positions increased the constitutive activity 5.7- and 2.3-fold, respectively, compared with EBI2 wild type (wt). In contrast, ArgII:20 (Arg(87)) in TM-2 acted as a positive regulator, as substitution to Ala, but not to Lys, decreased the constitutive activity more than 7-fold compared with wt EBI2. IleIII:03 (Ile106) is located only 4 angstrom from ArgII: 20, and a favorable electrostatic interaction with ArgII: 20 was created by introduction of Glu in III:03, given that the activity increased to 4.4-fold of that wt EBI2. It is noteworthy that swapping these charges by introduction of Glu in II:20 and Arg in III:03 resulted in a 2.7-fold increase compared with wt EBI2, thereby rescuing the two signaling-deficient single mutations, which exhibited a 3.8- to 4.5-fold decrease in constitutive activity. The uncovering of these molecular mechanisms for EBI2 activation is important from a drug development point of view, in that it may facilitate the rational design and development of small-molecule inverse agonists against EBI2 of putative importance as antiviral- or immune modulatory therapy.