Tunuguntla Ramya H, Escalada Artur, A Frolov Vadim, Noy Aleksandr
Biosciences and Biotechnology Division, Physics and Life Sciences Directorate, Lawrence Livermore National Laboratory, Livermore, California, USA.
Biofisika Institute (CSIC, UPV/EHU), University of the Basque Country, Leioa, Spain.
Nat Protoc. 2016 Oct;11(10):2029-2047. doi: 10.1038/nprot.2016.119. Epub 2016 Sep 22.
Carbon nanotube porins (CNTPs) are 10- to 20-nm-long segments of lipid-stabilized single-walled carbon nanotubes (CNTs) that can be inserted into phospholipid membranes to form nanometer-scale-diameter pores that approximate the geometry and many key transport characteristics of biological membrane channels. We describe protocols for CNTP synthesis by ultrasound-assisted cutting of long CNTs in the presence of lipid amphiphiles, and for validation of CNTP incorporation into a lipid membrane using a proton permeability assay. In addition, we describe protocols for measuring conductance of individual CNTPs in planar lipid bilayers and plasma membranes of live cells. The protocol for the preparation and testing of the CNTPs in vesicle systems takes 3 d, and single CNTP conductance measurements take 2-5 h. The CNTPs produced by this cutting protocol remain stable and active for at least 10-12 weeks.
碳纳米管孔蛋白（CNTPs）是脂质稳定的单壁碳纳米管（CNTs）10至20纳米长的片段，可插入磷脂膜中形成纳米级直径的孔，这些孔近似生物膜通道的几何形状和许多关键转运特性。我们描述了在脂质两亲物存在下通过超声辅助切割长碳纳米管来合成CNTPs的方案，以及使用质子渗透性测定法验证CNTPs掺入脂质膜的方案。此外，我们描述了测量平面脂质双层和活细胞质膜中单个CNTPs电导的方案。在囊泡系统中制备和测试CNTPs的方案需要3天，单个CNTPs电导测量需要2至5小时。通过这种切割方案产生的CNTPs至少在10至12周内保持稳定和活性。
