Functional map of arrestin-1 at single amino acid resolution
Functional map of arrestin-1 at single amino acid resolution
复制标题
DOI：
10.1073/pnas.1319402111
复制
发表时间：
2014-02-04
影响因子：
11.1
通讯作者：
Standfuss, Joerg
中科院分区：
文献类型：
--
作者：
Ostermaier, Martin K.;Peterhans, Christian;Standfuss, Joerg
抑制蛋白作为衔接蛋白发挥作用，介导G蛋白偶联受体（GPCR）脱敏、内化和额外的信号传导。在这里，我们比较了结合的GPCR视紫红质的403突变体的arrestin-1覆盖其完整的序列。这种全面和公正的诱变方法提供了一个功能层面的晶体结构的无活性，预活化p44和磷酸肽结合arrestins，并将指导我们的arrestin-GPCR复合物的理解。所提出的功能图定量地连接在极性核心和沿着C尾部的抑制蛋白的关键相互作用。一系列氨基酸（Phe 375、Phe 377、Phe 380和Arg 382）将C尾锚在阻断受体结合的位置。视紫红质C末端中的磷酸盐与Arg 29的相互作用控制C-尾交换机制，其中抑制蛋白的C尾被释放并暴露几个带电氨基酸（Lys 14、Lys 15、Arg 18、Lys 20、Lys 110和Lys 300）以结合磷酸化受体C末端。除了这种arrestin磷酸传感器，我们的数据揭示了手指（Gln 69和Asp 73-Met 75）和L249-S252和Y254）中的几个氨基酸补丁，可以作为直接结合界面。C结构域边缘的一段氨基酸（Trp 194-Ser 199、Gly 337-Gly 340、Thr 343和Thr 345）可以作为膜锚，第二个视紫红质的结合界面，或在复合物形成时更靠近中心环重排。我们讨论这些接口的背景下，实验指导的对接晶体结构之间的arrestin和光激活视紫红质。
Arrestins function as adapter proteins that mediate G protein-coupled receptor (GPCR) desensitization, internalization, and additional rounds of signaling. Here we have compared binding of the GPCR rhodopsin to 403 mutants of arrestin-1 covering its complete sequence. This comprehensive and unbiased mutagenesis approach provides a functional dimension to the crystal structures of inactive, preactivated p44 and phosphopeptide-bound arrestins and will guide our understanding of arrestin-GPCR complexes. The presented functional map quantitatively connects critical interactions in the polar core and along the C tail of arrestin. A series of amino acids (Phe375, Phe377, Phe380, and Arg382) anchor the C tail in a position that blocks binding of the receptor. Interaction of phosphates in the rhodopsin C terminus with Arg29 controls a C-tail exchange mechanism in which the C tail of arrestin is released and exposes several charged amino acids (Lys14, Lys15, Arg18, Lys20, Lys110, and Lys300) for binding of the phosphorylated receptor C terminus. In addition to this arrestin phosphosensor, our data reveal several patches of amino acids in the finger (Gln69 and Asp73-Met75) and the lariat loops (L249-S252 and Y254) that can act as direct binding interfaces. A stretch of amino acids at the edge of the C domain (Trp194-Ser199, Gly337-Gly340, Thr343, and Thr345) could act as membrane anchor, binding interface for a second rhodopsin, or rearrange closer to the central loops upon complex formation. We discuss these interfaces in the context of experimentally guided docking between the crystal structures of arrestin and light-activated rhodopsin.