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Imaging transmembrane dynamics of biomolecules at live cell plasma membranes using quenchers in extracellular environment

By Wenqing Hou, Dongfei Ma, Xiaolong He, Weijing Han, Jianbing Ma, Hao Wang, Chunhua Xu, Ruipei Xie, Qi-hui Fan, Fangfu Ye, Shuxin Hu, Ying Lu, Ming Li

Posted 31 Aug 2020
bioRxiv DOI: 10.1101/2020.08.30.266148

It is a big challenge to measure position changes of biomolecules in the direction normal to the plasma membranes of living cells. We developed a one donor-multiple quenchers Forster resonance energy transfer method by using non-fluorescent quenchers in the extracellular environment. It senses subnanometer position changes of a fluorophore-labeled biomolecule in the plasma membrane. The method was validated by monitoring flip-flops of individual lipid molecules incorporated in plasma membranes. We studies membrane perforation by a host defense peptide from the extracellular side and found that the pore-forming peptide is dynamic, switching among different insertion depths. The method is especially useful in studying interactions of membrane proteins with the inner surfaces of plasma membranes. Our method will find wide applications in systematic analysis of fundamental cellular processes at plasma membranes.

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