Specific heteromeric association of four transmembrane peptides derived from platelet glycoprotein Ib-IX complex.
Specific heteromeric association of four transmembrane peptides derived from platelet glycoprotein Ib-IX complex.
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DOI：
10.1016/j.jmb.2008.07.037
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发表时间：
2008-10-03
影响因子：
5.6
通讯作者：
Li, Renhao
中科院分区：
文献类型：
--
作者：
Luo, Shi-Zhong;Li, Renhao
关键词：
糖蛋白（GP）Ib-IX复合物作为血管性血友病因子（vonWillebrand factor，vWF）的血小板表面受体，在止血和血栓形成中起重要作用。该复合物如何由GP Ibα、GP Ibβ和GP IX亚基（均为I型跨膜蛋白）组装而成，目前尚不完全清楚。遗传和突变分析已经确定了这些亚基的跨膜（TM）结构域作为复杂组装的积极参与者。在这项研究中，含有每个亚基的跨膜结构域的肽已被生产和他们的相互作用的特点。在SDS电泳和TOXCAT试验中，只有Ibβ TM序列，而不是Ibα和IX对应序列，可以形成同源寡聚体。我们先前观察到，在洗涤剂胶束中，可以从分离的Ibα和Ibβ TM肽产生通过天然质膜二硫键连接的Ibβ-Ibα-Ibβ肽复合物（αβ2），在此我们表明，添加IX TM肽有助于天然αβ2复合物的形成，在表达GP Ib-IX复合物的细胞中重现IX亚基的相同作用。在胶束中，供体标记的αβ2肽复合物与受体标记的IX TM肽之间存在特异性荧光共振能量转移。最后，IX TM肽中的突变D135 K可阻碍αβ2复合物形成和能量转移，与其在全长复合物中的报告效应一致。总之，我们的结果直接证明了分离的Ibα、Ibβ和IX TM肽之间的天然异聚体相互作用，这为GP Ib-IX复合物中TM结构域的四螺旋束模型提供了支持，并为进一步的结构分析铺平了道路。本研究中开发的方法也可适用于多个TM螺旋之间的异聚体相互作用的其他研究。
As the receptor on platelet surface for von Willebrand factor, glycoprotein (GP) Ib-IX complex is critically involved in hemostasis and thrombosis. How the complex is assembled from GP Ibα, GP Ibβ and GP IX subunits, all of which are type I transmembrane proteins, is not entirely clear. Genetic and mutational analyses have identified the transmembrane (TM) domains of these subunits as active participants in complex assembly. In this study, peptides containing the transmembrane domain of each subunit have been produced and their interaction with one another characterized. Only the Ibβ TM sequence, not Ibα and IX counterparts, can form homo-oligomers in SDS electrophoresis and TOXCAT assays. Following up on our earlier observation that a Ibβ-Ibα-Ibβ peptide complex (αβ2) linked through native juxtamembrane disulfide bonds could be produced from isolated Ibα and Ibβ TM peptides in detergent micelles, here we show that addition of the IX TM peptide facilitates formation of the native αβ2 complex, reproducing the same effect by the IX subunit in cells expressing GP Ib-IX complex. Specific fluorescence resonance energy transfer was observed between donor-labeled αβ2 peptide complex and acceptor-conjugated IX TM peptide in micelles. Finally, the mutation D135K in the IX TM peptide could hamper both the αβ2 complex formation and the energy transfer, consistent with its reported effect in the full-length complex. Overall, our results have demonstrated directly the native-like heteromeric interaction among the isolated Ibα, Ibβ and IX TM peptides, which provides support for the four-helical bundle model of the TM domains in GP Ib-IX complex and paves the way for further structural analysis. The methods developed in this study may also be applicable to other studies of heteromeric interaction among multiple TM helices.
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影响因子：
4.8
作者：
Gurezka, R;Langosch, D
通讯作者：
Langosch, D
DOI：
10.1073/pnas.78.5.2712
发表时间：
1981-01-01
期刊：
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA-BIOLOGICAL SCIENCES
影响因子：
--
作者：
CLEMETSON, KJ;NAIM, HY;LUSCHER, EF
通讯作者：
LUSCHER, EF
影响因子：
4.8
作者：
Bodnar, RJ;Xi, XD;Du, XP
通讯作者：
Du, XP
影响因子：
6.7
作者：
González-Manchón, C;Butta, N;Parrilla, R
通讯作者：
Parrilla, R
影响因子：
20.3
作者：
Kahn, ML;Diacovo, TG;Coughlin, S
通讯作者：
Coughlin, S