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首页> 外文会议>Conference on Optical Diagnostics of Living Cells Ⅴ, Jan 23-25, 2002, San Jose, USA >Investigating Intermolecular Forces associated with Thrombus Initiation Using Optical Tweezers
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Investigating Intermolecular Forces associated with Thrombus Initiation Using Optical Tweezers

机译：使用光镊研究与血栓形成相关的分子间力

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摘要

Thrombus formation occurs when a platelet membrane receptor, glycoprotein (GP) Ib-IX-V complex, binds to its ligand, von Willebrand factor (vWf), in the subendothelium or plasma. To determine which GP Ib-IX-V amino acid sequences are critical for bond formation, we have used optical tweezers to measure forces involved in the binding of vWf to GP Ib-IX-V variants. Inasmuch as GP Ibα subunit is the primary component in human GP Ib-IX-V complex that binds to vWf, and that canine GP Ibα, on the other hand, does not bind to human vWf, we progressively replaced human GP Ibα amino acid sequences with canine GP Ibα sequences to determine the sequences essential for vWf/GP Ibα binding. After measuring the adhesive forces between optically trapped, vWf-coated beads and GP Ibα variants expressed on mammalian cells, we determined that leucine-rich repeat 2 of GP Ibα was necessary for vWf/GP Ib-IX-V bond formation. We also found that deletion of the N-terminal flanking sequence and leucine-rich repeat 1 reduced adhesion strength to vWf but did not abolish binding. While divalent cations are known to influence binding of vWf, addition of 1mM CaCl_2 had no effect on measured vWf/GP Ibα bond strengths.
机译：当血小板膜受体糖蛋白（GP）Ib-IX-V复合物在内皮下或血浆中与其配体von Willebrand因子（vWf）结合时，就会形成血栓形成。为了确定哪些GP Ib-IX-V氨基酸序列对于键形成至关重要，我们使用了光镊来测量vvf与GP Ib-IX-V变体结合中所涉及的力。由于GPIbα亚基是人GP Ib-IX-V复合物中与vWf结合的主要成分，而犬GPIbα不与人vWf结合，因此我们逐步取代了人GPIbα氨基酸序列用犬GPIbα序列确定vWf / GPIbα结合必不可少的序列。在测量光学捕获的，vWf包被的珠子与在哺乳动物细胞上表达的GPIbα变体之间的粘附力后，我们确定GPIbα的富含亮氨酸的重复序列2对于vWf / GP Ib-IX-V键形成是必需的。我们还发现删除N末端侧翼序列和富含亮氨酸的重复序列1降低了对vWf的粘附强度，但并未消除结合。尽管已知二价阳离子会影响vWf的结合，但添加1mM CaCl_2对测得的vWf / GPIbα键强度没有影响。

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