Department of Chemical Engineering, Polytechnique Montreal, Montreal, QC, H3T 1J4, Canada.
Human Health Therapeutics Research Centre, National Research Council of Canada, Montreal, QC, H4P 2R2, Canada.
Sci Rep. 2022 Jul 7;12(1):11520. doi: 10.1038/s41598-022-15215-5.
Several key mutations in the Spike protein receptor binding domain (RBD) have been identified to influence its affinity for the human Angiotensin-Converting Enzyme 2 (ACE2). Here, we perform a comparative study of the ACE2 binding to the wild type (Wuhan) RBD and some of its variants: Alpha B.1.1.7, Beta B.1.351, Delta B.1.617.2, Kappa B.1.617.1, B.1.1.7 + L452R and Omicron B.1.1.529. Using a coiled-coil mediated tethering approach of ACE2 in a novel surface plasmon resonance (SPR)-based assay, we measured interactions at different temperatures. Binding experiments at 10 °C enhanced the kinetic dissimilarities between the RBD variants and allowed a proper fit to a Langmuir 1:1 model with high accuracy and reproducibility, thus unraveling subtle differences within RBD mutants and ACE2 glycovariants. Our study emphasizes the importance of SPR-based assay parameters in the acquisition of biologically relevant data and offers a powerful tool to deepen our understanding of the role of the various RBD mutations in ACE2 interaction binding parameters.
一些关键的刺突蛋白受体结合域（RBD）突变已被确定会影响其与人类血管紧张素转换酶 2（ACE2）的亲和力。在这里，我们对 ACE2 与野生型（武汉）RBD 及其一些变体的结合进行了比较研究：Alpha B.1.1.7、Beta B.1.351、Delta B.1.617.2、Kappa B.1.617.1、B.1.1.7 + L452R 和 Omicron B.1.1.529。我们使用一种新型表面等离子体共振（SPR）基于测定的 ACE2 卷曲螺旋介导的系绳方法，在不同温度下测量相互作用。在 10°C 下进行的结合实验增强了 RBD 变体之间的动力学差异，并允许对高准确性和重现性的 Langmuir 1:1 模型进行适当拟合，从而揭示了 RBD 突变体和 ACE2 糖型变体内部的细微差异。我们的研究强调了基于 SPR 的测定参数在获得具有生物学相关性数据方面的重要性，并提供了一种强大的工具，可以加深我们对 ACE2 相互作用结合参数中各种 RBD 突变的作用的理解。
