Force spectroscopy of the leukocyte function-associated antigen-1/intercellular adhesion molecule-1 interaction.
Force spectroscopy of the leukocyte function-associated antigen-1/intercellular adhesion molecule-1 interaction.
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DOI：
10.1016/s0006-3495(02)73987-8
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发表时间：
2002-10
影响因子：
3.4
通讯作者：
Xiaohui Zhang;E. Wojcikiewicz;V. Moy
中科院分区：
文献类型：
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作者：
Xiaohui Zhang;E. Wojcikiewicz;V. Moy
白细胞功能相关抗原-1（LFA-1）与其同源配体细胞间粘附分子-1（ICAM-1）之间的相互作用在白细胞粘附中起着至关重要的作用。由于细胞及其粘附组分受到来自周围血流的外部扰动，因此重要的是要了解稳态和存在外部拉力时LFA-1/ICAM-1相互作用的结合特性。在这里，我们报告原子力显微镜（AFM）测量的LFA-1从ICAM-1的解结合。单分子测量揭示了对应于LFA-1/ICAM-1复合物解离的能量景观，并为定义平衡时和外力影响下的复合物的能量决定因素提供了基础。AFM力测量在实验系统中进行，该实验系统由附着于AFM悬臂末端的表达LFA-1的T细胞杂交瘤3A 9和表达ICAM-1的并置表面组成。在覆盖力加载速率的三个数量级变化的测量中，LFA-1/ICAM-1力谱（即，松解力与加载速率）揭示了快速和缓慢加载状态，其特征分别是陡峭的内部活化势垒和宽的外部活化势垒。Mg 2+的加入，一种稳定LFA-1/ICAM-1相互作用的辅因子，提高了复合物在缓慢加载方案中的解结合力。相反，EDTA的存在抑制了LFA-1/ICAM-1复合物的内屏障。这些结果表明，LFA-1/ICAM-1相互作用的平衡解离常数是由复合物的外部活化势垒的能量学调节的，而复合物抵抗拉力的能力是由二价阳离子依赖性的内部活化势垒决定的。
Interactions between leukocyte function-associated antigen-1 (LFA-1) with its cognate ligand, intercellular adhesion molecule-1 (ICAM-1) play a crucial role in leukocyte adhesion. Because the cell and its adhesive components are subject to external perturbation from the surrounding flow of blood, it is important to understand the binding properties of the LFA-1/ICAM-1 interaction in both steady state and in the presence of an external pulling force. Here we report on atomic force microscopy (AFM) measurements of the unbinding of LFA-1 from ICAM-1. The single molecule measurements revealed the energy landscape corresponding to the dissociation of the LFA-1/ICAM-1 complex and provided the basis for defining the energetic determinants of the complex at equilibrium and under the influence of an external force. The AFM force measurements were performed in an experimental system consisting of an LFA-1-expressing T cell hybridoma, 3A9, attached to the end of the AFM cantilever and an apposing surface expressing ICAM-1. In measurements covering three orders of magnitude change in force loading rate, the LFA-1/ICAM-1 force spectrum (i.e., unbinding force versus loading rate) revealed a fast and a slow loading regime that characterized a steep inner activation barrier and a wide outer activation barrier, respectively. The addition of Mg2+, a cofactor that stabilizes the LFA-1/ICAM-1 interaction, elevated the unbinding force of the complex in the slow loading regime. In contrast, the presence of EDTA suppressed the inner barrier of the LFA-1/ICAM-1 complex. These results suggest that the equilibrium dissociation constant of the LFA-1/ICAM-1 interaction is regulated by the energetics of the outer activation barrier of the complex, while the ability of the complex to resist a pulling force is determined by the divalent cation-dependent inner activation barrier.