Ligand affinity of the 67-kD elastin/laminin binding protein is modulated by the protein's lectin domain: visualization of elastin/laminin-receptor complexes with gold-tagged ligands.
Ligand affinity of the 67-kD elastin/laminin binding protein is modulated by the protein's lectin domain: visualization of elastin/laminin-receptor complexes with gold-tagged ligands.
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DOI：
10.1083/jcb.113.1.187
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发表时间：
1991-04
期刊：
影响因子：
--
通讯作者：
Sheetz MP
中科院分区：
文献类型：
--
作者：
Mecham RP;Whitehouse L;Hay M;Hinek A;Sheetz MP
使用视频增强显微镜检查弹性蛋白或层粘连蛋白包被的金颗粒与活细胞表面弹性蛋白结合蛋白的相互作用。通过实时观察结合事件，可以确定配体结合的特异性和亲合力，并分析受体-配体复合物在质膜平面中的运动。尽管由于颗粒和细胞表面之间可能存在多种相互作用，因此很难严格解释结合和释放的速率，但结合的相对变化揭示了配体-受体原位相互作用的亲和力调节的重要方面。研究发现弹性蛋白和层粘连蛋白都会竞争与细胞表面的结合，并且乳糖显着降低了弹性蛋白和层粘连蛋白受体的亲和力。这些发现得到了去污剂溶解受体的体外研究的支持。此外，通过与细胞骨架结合来固定配体-受体复合物，显着降低了结合颗粒离开受体的能力。配体包被的金与活细胞结合的动力学变化表明，层粘连蛋白和弹性蛋白的结合均受到乳糖的抑制，并且受体与细胞骨架的附着增加了其与配体的亲和力。
Video-enhanced microscopy was used to examine the interaction of elastin- or laminin-coated gold particles with elastin binding proteins on the surface of live cells. By visualizing the binding events in real time, it was possible to determine the specificity and avidity of ligand binding as well as to analyze the motion of the receptor-ligand complex in the plane of the plasma membrane. Although it was difficult to interpret the rates of binding and release rigorously because of the possibility for multiple interactions between particles and the cell surface, relative changes in binding have revealed important aspects of the regulation of affinity of ligand-receptor interaction in situ. Both elastin and laminin were found to compete for binding to the cell surface and lactose dramatically decreased the affinity of the receptor(s) for both elastin and laminin. These findings were supported by in vitro studies of the detergent-solubilized receptor. Further, immobilization of the ligand-receptor complexes through binding to the cytoskeleton dramatically decreased the ability of bound particles to leave the receptor. The changes in the kinetics of ligand-coated gold binding to living cells suggest that both laminin and elastin binding is inhibited by lactose and that attachment of receptor to the cytoskeleton increases its affinity for the ligand.