A practical guide for fast implementation of SNARE-mediated liposome fusion

作     者：Shen Wang Cong Ma 

作者机构：Key Laboratory of Molecular Biophysics of the Ministry of Education College of Life Science and TechnologyHuazhong University of Science and Technology 

出 版 物：《Biophysics Reports》 (生物物理学报)

年 卷 期：2024年第10卷第1期

页      面：31-40页

核心收录：

学科分类：08[工学] 0836[工学-生物工程] 

基　　金：supported by the National Funds for Distinguished Young Scholars of China (32225024) the National Natural Science Foundation of China (92254302 and 32071224) the National Science and Technology Major Project of China (2021ZD0202501) 

主　　题：Liposome fusion SNARE In-vitro reconstitution Protocol 

摘      要：Soluble N-ethylmaleimide-sensitive factor attachment protein receptor(SNAER) family proteins are the engines of most intra-cellular and exocytotic membrane fusion pathways(Jahn and Scheller 2006).Over the past two decades, in-vitro liposome fusion has been proven to be a powerful tool to reconstruct physiological SNARE-mediated membrane fusion processes(Liu et al. 2017). The reconstitution of the membrane fusion process not only provides direct evidence of the capability of the cognate SNARE complex in driving membrane fusion but also allows researchers to study the functional mechanisms of regulatory proteins in related pathways(Wickner and Rizo 2017). Heretofore, a variety of delicate methods for in-vitro SNARE-mediated liposome fusion have been established(Bao et al. 2018;Diao et al. 2012; Duzgunes 2003; Gong et al. 2015; Heo et al. 2021; Kiessling et al. 2015; Kreye et al.2008; Kyoung et al. 2013; Liu et al. 2017; Scott et al. 2003). Although technological advances have made reconstitution more physiologically relevant, increasingly elaborate experimental procedures, instruments, and data processing algorithms nevertheless hinder the non-experts from setting up basic SNARE-mediated liposome fusion assays. Here, we describe a low-cost, timesaving, and easy-to-handle protocol to set up a foundational in-vitro SNARE-mediated liposome fusion assay based on our previous publications(Liu et al. 2023; Wang and Ma 2022). The protocol can be readily adapted to assess various types of SNARE-mediated membrane fusion and the actions of fusion regulators by using appropriate alternative additives(e.g., proteins, macromolecules, chemicals, etc.). The total time required for one round of the assay is typically two days and could be extremely compressed into one day.