通过时间分辨荧光共振能量转移（TR-FRET）检测活细胞中 SARS-CoV-2 刺突蛋白与 ACE2 的结合。

Detection of SARS-CoV-2 spike protein binding to ACE2 in living cells by TR-FRET.

机构信息

Université de Paris, Institut Cochin, INSERM, CNRS, 75014 Paris, France.

出版信息

STAR Protoc. 2022 Mar 18;3(1):101024. doi: 10.1016/j.xpro.2021.101024. Epub 2021 Nov 22.

DOI:10.1016/j.xpro.2021.101024

PMID:34841271

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC8606263/

Abstract

The SARS-CoV-2 coronavirus infects human cells through the interaction of the viral envelope spike protein (IPR044366) with the human angiotensin-converting enzyme 2 (ACE2), expressed at the surface of target cells. Here, we describe a detailed protocol to measure the binding of the receptor binding domain (RBD) of spike to ACE2 by time-resolved fluorescence resonance energy transfer (TR-FRET). The assay detects the spike/ACE2 interaction in physiologically relevant cellular contexts and is suitable for high-throughput investigation of interfering small-molecule compounds and antibodies. For complete details on the use and execution of this protocol, please refer to Cecon et al. (2021).

摘要

新型冠状病毒通过其包膜刺突蛋白（IPR044366）与在靶细胞表面表达的人血管紧张素转化酶 2（ACE2）的相互作用感染人类细胞。在这里，我们描述了一种通过时间分辨荧光共振能量转移（TR-FRET）测量受体结合域（RBD）与 ACE2 结合的详细方案。该测定法在生理相关的细胞环境中检测刺突/ACE2 相互作用，适用于干扰小分子化合物和抗体的高通量研究。有关该方案使用和执行的完整详细信息，请参阅 Cecon 等人。（2021）。