Department of Cell Biology, Yale University School of Medicine, New Haven, Connecticut.
Department of Cell Biology, Yale University School of Medicine, New Haven, Connecticut; Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut.
Biophys J. 2021 Dec 21;120(24):5454-5465. doi: 10.1016/j.bpj.2021.11.2884. Epub 2021 Nov 20.
Despite their wide applications in soluble macromolecules, optical tweezers have rarely been used to characterize the dynamics of membrane proteins, mainly due to the lack of model membranes compatible with optical trapping. Here, we examined optical trapping and mechanical properties of two potential model membranes, giant and small unilamellar vesicles (GUVs and SUVs, respectively) for studies of membrane protein dynamics. We found that optical tweezers can stably trap GUVs containing iodixanol with controlled membrane tension. The trapped GUVs with high membrane tension can serve as a force sensor to accurately detect reversible folding of a DNA hairpin or membrane binding of synaptotagmin-1 C2AB domain attached to the GUV. We also observed that SUVs are rigid enough to resist large pulling forces and are suitable for detecting protein conformational changes induced by force. Our methodologies may facilitate single-molecule manipulation studies of membrane proteins using optical tweezers.
尽管光镊在可溶性大分子中有着广泛的应用，但由于缺乏与光阱兼容的模型膜，光镊很少用于表征膜蛋白的动力学。在这里，我们研究了两种潜在的模型膜，即巨大和小单层囊泡（GUV 和 SUV，分别），用于研究膜蛋白动力学。我们发现，光镊可以稳定地捕获含有碘海醇的 GUV，并控制其膜张力。具有高膜张力的捕获 GUV 可以作为力传感器，准确检测 DNA 发夹的可逆折叠或与 GUV 相连的突触融合蛋白-1 C2AB 结构域的膜结合。我们还观察到，SUV 足够坚固，可以抵抗较大的拉力，适合检测力诱导的蛋白质构象变化。我们的方法学可能有助于使用光镊进行膜蛋白的单分子操纵研究。
