Biological Membranes and Atomic Force Microscopy

生物膜和相关系统正在研究通过原子力显微镜（AFM）和其他生物物理技术在这个项目包含几个合作。(1)我们继续与B博士合作。J. Litman和S.- L. Niu（NIAAA，NIH）使用原子力显微镜（AFM）以亚纳米分辨率表征视紫红质（G蛋白偶联受体（GPCR））在视觉通路的天然视杆外段膜中以及用二棕榈酰磷脂酰胆碱（DPPC，di 16：0 PC）和多不饱和脂质（如DDPC（di-22：6 n-3 PC））重建的膜中的结构和功能。我们已经获得了新的AFM数据在更广泛的范围内的原生视觉磁盘和重组视紫红质/膜系统在一组更广泛的条件。我们开发了新的AFM成像分析方法和生物物理模拟，以进一步揭示视紫红质分子的组织，并探讨视紫红质信号和脂质膜环境之间的联系。(2)我们与Ralph Nossal博士（NICHD，NIH）和Eileen M. Lafer和K.普拉萨德（德克萨斯大学，圣安东尼奥）。CCV是重要的生物膜和多蛋白复合物，其在受体介导的内吞作用和从trans-Golgi网络的细胞内运输中起核心作用。我们使用AFM和专用的生物物理数据分析方法测量了CCV的机械性能。我们已经证实了已知的拓扑网格蛋白外壳结构从电子显微镜和进一步移动到产生CCV变形作为定量响应的压缩力在100 pN范围内。将CCV建模为复合弹性薄壳，我们估计完整CCV的弯曲刚度约为其构成外部网格蛋白多面体晶格和内部磷脂膜的10倍。这表明天然CCV中的衔接蛋白层在其网格蛋白外壳与装载有受体和配体的内脂质膜之间提供部分偶联。(3)我们继续使用AFM，生物物理数据分析，纳米技术和其他科学方法进行生物膜和相关研究。

Biological membranes and related systems are being investigated via atomic force microscopy (AFM) and other biophysical technologies in this project containing several collaborations. (1) We continue our collaboration with Drs. B. J. Litman and S.-L. Niu (NIAAA, NIH), using atomic force microscopy (AFM) to characterize at sub-nanometer resolutions structure and function of the protein rhodopsin, a G-protein coupled receptor (GPCR), in native rod outer segment membranes of the vision pathway and in reconstituted membranes with dipalmitoylphosphatidylcholine (DPPC, di16:0PC) and polyunsatuatrated lipids such as DDPC (di-22:6n-3PC). We have obtained new AFM data on a broader range of native vision disks and reconstituted rhodopsin/membrane system over a broader set of conditions. We have developed new AFM imaging analysis methods and biophysical simulations to further reveal rhodopsin molecule organization and to explore the connection between rhodopsin signaling and the lipid membrane environment. (2) We have advanced our cutting edge AFM studies of clathrin coated vesicles (CCVs) in collaboration with Dr. Ralph Nossal (NICHD, NIH) and Drs. Eileen M. Lafer and K. Prasad (University Texas, San Antonio). CCVs are important biological membrane and multi-protein complexes that play a central role in receptor-mediated endocytosis and intracellular trafficking from the trans-Golgi network. We have measured the mechanical properties of CCVs using AFM and dedicated biophysical data analysis methods. We have confirmed the topological clathrin coat structures known from electron microscopy and moved further to yield CCV deformation as a quantitative response to the compression force in the 100 pN range. Modeling CCVs as composite elastic thin shells, we estimate that the bending rigidity of intact CCVs is about 10 times that of its constituting outer clathrin polyhedral lattice and inner phospholipids membrane. This suggests that the adaptor protein layer in native CCVs provides a partial coupling between its clathrin coat and the inner lipid membrane loaded with receptors and ligands. (3) We continue our biological membrane and related studies using AFM, biophysical data analyses, nanotechnology, and other scientific methods.