Cellular life relies on membranes, which provide a resilient and adaptive cell boundary. Many essential processes depend upon the ease with which the membrane is able to deform and bend, features that can be characterized by the bending rigidity. Quantitative investigations of such mechanical properties of biological membranes have primarily been undertaken in solely lipid bilayers and frequently in the absence of buffers. In contrast, much less is known about the influence of integral membrane proteins on bending rigidity under physiological conditions. We focus on an exemplar member of the ubiquitous major facilitator superfamily of transporters and assess the influence of lactose permease on the bending rigidity of lipid bilayers. Fluctuation analysis of giant unilamellar vesicles (GUVs) is a useful means to measure bending rigidity. We find that using a hydrogel substrate produces GUVs that are well suited to fluctuation analysis. Moreover, the hydrogel method is amenable to both physiological salt concentrations and anionic lipids, which are important to mimic key aspects of the native lactose permease membrane. Varying the fraction of the anionic lipid in the lipid mixture DOPC/DOPE/DOPG allows us to assess the dependence of membrane bending rigidity on the topology and concentration of an integral membrane protein in the lipid bilayer of GUVs. The bending rigidity gradually increases with the incorporation of lactose permease, but there is no further increase with greater amounts of the protein in the membrane.

细胞生命依赖于膜，膜提供了有弹性和适应性的细胞边界。许多基本过程取决于膜变形和弯曲的难易程度，这些特征可以通过弯曲刚度来表征。对生物膜的这种机械性能的定量研究主要是在单独的脂质双层中进行的，并且经常在没有缓冲剂的情况下进行。相比之下，人们对生理条件下完整膜蛋白对弯曲刚度的影响知之甚少。我们关注普遍存在的转运蛋白主要促进子超家族的典型成员，并评估乳糖通透酶对脂质双层弯曲刚度的影响。巨型单层囊泡（GUV）的波动分析是测量弯曲刚度的有用方法。我们发现使用水凝胶基质产生的 GUV 非常适合波动分析。此外，水凝胶方法适用于生理盐浓度和阴离子脂质，这对于模拟天然乳糖渗透酶膜的关键方面非常重要。改变脂质混合物 DOPC/DOPE/DOPG 中阴离子脂质的比例，使我们能够评估膜弯曲刚度对 GUV 脂质双层中完整膜蛋白的拓扑和浓度的依赖性。随着乳糖通透酶的掺入，弯曲刚度逐渐增加，但随着膜中蛋白质含量的增加，弯曲刚度不再进一步增加。