Paratope and epitope mapping of the antithrombotic antibody 6B4 in complex with platelet glycoprotein Ibα
Paratope and epitope mapping of the antithrombotic antibody 6B4 in complex with platelet glycoprotein Ibα
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DOI：
10.1074/jbc.m701826200
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发表时间：
2007-08-10
影响因子：
4.8
通讯作者：
Deckmyn, Hans
中科院分区：
文献类型：
--
作者：
Fontayne, Alexandre;De Maeyer, Bauke;Deckmyn, Hans
单克隆抗体6 B4通过与糖蛋白Ib α（GPIb α）的柔性环（也称为β开关区（氨基酸230-242））结合，在非人灵长类动物中具有强效抗血栓形成作用。在高剪切应力条件下，这种相互作用阻断GPIb α和血管性血友病因子之间的特异性相互作用，抑制血小板沉积到受损的血管壁，这是动脉血栓形成发病机制中的关键事件。为了了解这种抗体和它的抗原之间的相互作用在氨基酸水平上，我们在这里报告的6 B4和GPIb α，分别通过使用计算机建模和定点诱变的互补位和表位的识别。使用对接程序ZDOCK（刚体对接）和HADDOCK（柔性对接）来模拟6 B4与GPIb α突变体的相互作用，并分别测定它们结合GPIb α和6 B4的能力。根据这些数据，发现6 B4的互补位主要由5个残基形成：分别位于轻链CDR 1和-3的Tyr（27 D）、Lys（27 E）、Asp（28）和Glu（93），以及重链CDR 3的Tyr（100 C）。这些残基形成谷，其中GPIb α柔性环可以通过残基Asp（235）和Lys（237）结合。最后利用实验结果建立了更精确的对接模型。两者合计，这些信息提供了新的衍生的抗血栓性质的先导化合物的设计准则。
The monoclonal antibody 6B4 has a potent antithrombotic effect in nonhuman primates by binding to the flexible loop, also known as the beta-switch region ( amino acids 230-242), of glycoprotein Ib alpha(GPIb alpha). This interaction blocks, in high shear stress conditions, the specific interaction between GPIb alpha and von Willebrand factor suppressing platelet deposition to the damaged vessel wall, a key event in the pathogenesis of arterial thrombosis. To understand the interactions between this antibody and its antigen at the amino acid level, we here report the identification of the paratope and epitope in 6B4 and GPIb alpha, respectively, by using computer modeling and site-directed mutagenesis. The docking programs ZDOCK ( rigid body docking) and HADDOCK ( flexible docking) were used to model the interaction of 6B4 with GPIb alpha mutants were constructed and assayed for their capacity to bind GPIb alpha and 6B4, respectively. From these data, it is found that the paratope of 6B4 is mainly formed by five residues: Tyr(27D), Lys(27E), Asp(28), and Glu(93) located in light chain CDR1and -3, respectively, and Tyr(100C) of the heavy chain CDR3. These residues form a valley, where the GPIb alpha flexible loop can bind via residues Asp(235) and Lys(237). The experimental results were finally used to build a more accurate docking model. Taken together, this information provides guidelines for the design of new derivatized lead compounds with antithrombotic properties.