Binding of integrin alphaIIbbeta3 (αiibβ3) to its ligands is a highly restricted and regulated mechanism. Any modification of the protein structure yields a dysfunctional role, especially in a redox environment. Here, we examine the effect of nitrosative stress on the αiibβ3 reconstituted into nanodiscs. Using single molecule force spectroscopy, we measured the interaction between αiibβ3 and its ligand RGD and found that in the presence of exogenous nitric oxide (NO) two force regimes are generated: a low force regime of ~100pN indicating the presence of integrin in a normal status, and a broad spectrum of high force regime (~210-450pN) suggesting the protein modification/aggregation. By high resolution atomic force microscopy imaging, we demonstrate that both NO and nitrite (a stable product formed from NO) are involved in destabilizing the transmembrane protein complex leading to release of αiibβ3 from the lipid bilayer and protein aggregation. Our experimental setup opens new ways for testing in a membrane environment the effect of radical species on integrins under clinically relevant conditions.

中文翻译：

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亚硝化胁迫影响整联蛋白alphaIIbbeta3与它的配体的相互作用。

整联蛋白αIIbbeta3（αiibβ3）与其配体的结合是高度受限制和调节的机制。蛋白质结构的任何修饰都会产生功能障碍，尤其是在氧化还原环境中。在这里，我们研究了亚硝化应激对重构为纳米圆盘的αiibβ3的影响。使用单分子力谱法，我们测量了αiibβ3及其配体RGD之间的相互作用，发现在存在外源性一氧化氮（NO）的情况下，会产生两种力态：〜100pN的低力态表明正常情况下存在整联蛋白状态，以及广泛的高压模式（〜210-450pN），提示蛋白质修饰/聚集。通过高分辨率原子力显微镜成像，我们证明，NO和亚硝酸盐（由NO形成的稳定产物）均与稳定跨膜蛋白复合物有关，导致脂质双层中αiibβ3的释放和蛋白质聚集。我们的实验装置为在膜环境中测试自由基物质在临床相关条件下对整联蛋白的作用开辟了新途径。