Membrane Biophysics Via Atomic Force Microscopy

在这个项目中，原子力显微镜(AFM)正在与NIAID的合作者一起研究脂膜的物理化学性质和异质性，这对许多生物过程都很重要。利用环境控制室中的敲击模式AFM图像，在不同温度下解析了2-二肉豆蔻-sn-甘油-磷胆碱(DMPC)的主要相变的定量特征。这些AFM图像显示，在预期的DMPC链熔化温度附近有两个膜相。我们已经从精确的AFM厚度测量中量化了DMPC膜在链熔化时的显著减薄和机械软化。我们还构建了一个新的生物物理模型，该模型允许从AFM获得的依赖于温度的相分布估计热力学转变热、熵和膜相区大小。推导出的本征微区直径约为4.2 nm。DMPC膜的工作已经扩展到研究更多的生物膜微域，即(1)二棕榈酰磷脂酰胆碱(DPPC)、二月桂酰磷脂酰胆碱(DLPC)和胆固醇(Chol)的三脂混合物，以及(2)红细胞膜。我们发现，三脂膜在云母基质上形成了微米和纳米结构域；胆固醇水平和温度都影响这些结构域的大小和动态。我们开发了新的视频显微镜和图像分析来表征恶性疟原虫感染过程中的红细胞闪烁和边缘抖动现象。我们发现，寄生虫感染显著改变了细胞膜的动力学，在微循环中可能与疟疾的发病机制有关。总体而言，原子力显微镜和相关技术正在进一步发展，以阐明与膜相关的生物分子事件，这些事件对医学上的重要过程至关重要。

The physical-chemical properties and heterogeneities of lipid membranes, which are important to many biological processes, are being investigated by atomic force microscopy (AFM) in this project with NIAID collaborators. Quantitative features of the main phase transition in 2-dimyristoyl-sn-glycero-phosphocholine (DMPC) have been resolved by tapping-mode AFM images in an environmentally controlled chamber at various temperatures. These AFM images reveal two membrane phases near the expected DMPC chain-melting temperature. We have quantified the marked thinning and mechanical softening of the DMPC membrane upon chain melting from precise AFM thickness measurements. We have also constructed a novel biophysical model, which permits an estimate of the thermodynamic transition enthalpy, entropy, and the membrane phase domain size from our AFM-acquired temperature-dependent phase distributions. The deduced intrinsic domain size is about 4.2 nm in diameter. The work on DMPC membrane has been extended to examine the more biological membrane microdomains, or rafts, in (1) tri-lipid mixtures of dipalmitoyl phosphatidylcholine (DPPC), dilauroyl phosphatidylcholine (DLPC), and cholesterol (Chol), and (2) red blood cell membranes. We found that the tri-lipid membrane formed both microscopic and nanoscopic domains over mica substrates; both the cholesterol level and the temperature affected the sizes and dynamic of these domains. We developed new video microscopy and image analyses to characterize the red blood cell flicker and edge dithering phenomena during the Plasmodium falciparum malaria infection process. We found that the parasitic infection markedly modifies cell membrane dynamics, in potential relevance to malaria disease mechanism in microcirculations. Overall, the AFM and related technology are being advanced further to elucidate membrane-associated biomolecular events critical to medically important processes.