Connecting plasma membrane function to lipid structure and organization with asym

通过不对称将质膜功能与脂质结构和组织联系起来

基本信息

批准号：
8708115
负责人：
NOAH MALMSTADT
金额：
$ 29.14万
依托单位：
UNIVERSITY OF SOUTHERN CALIFORNIA
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-09-30 至 2016-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8708115
关键词：
Affect Automobile Driving Basic Science Behavior Binding Biochemistry Biological Biological Process Biotechnology Carbohydrates Cell membrane Cells Cellular Morphology Cellular Structures Charge Chemicals Cilia Complex Crowding Cytoplasm DNA-Protein Interaction Data Development Diabetes Mellitus Disease Drug Delivery Systems Drug Transport Extracellular Space Fluorescence Fluorescence Microscopy Gene Expression Genetic Transcription Geometry Health Human Individual Integral Membrane Protein Ions Lipid Bilayers Lipids Location Measures Mechanics Membrane Membrane Proteins Microfluidics Modeling Molecular Molecular Biology Molecular Medicine Molecular Structure Monitor Oligonucleotides Peptides Pharmaceutical Preparations Phosphatidylserines Physiological Processes Play Process Property Proteins Research Resistance Role Route Signal Transduction Site Structure Structure-Activity Relationship System Techniques Testing Toxic Environmental Substances Translations Transmembrane Domain Vesicle Viral Virus annexin A5 base cancer therapy cell behavior design interest lipid structure molecular asymmetry nucleic acid structure particle passive transport protein function public health relevance receptor research study scaffold small molecule solute tool unilamellar vesicle

项目摘要

DESCRIPTION (provided by applicant): Over the past half century, as molecular biology and biochemistry have developed to inform our understanding of disease, and as this understanding has driven the search for treatments in biotechnology and molecular medicine, the dominant theoretical scaffold for interpreting molecular behavior has been the structure-function relationship. Our understanding of molecular biology's structure-function relationships is largely limited, however, to the molecules of the central dogma: proteins and oligonucleotides. The vast variety of molecular structure outside of the central dogma, particularly among lipids and carbohydrates, suggests that these, too, can be understood in terms of structure driving function. In particular, there has been intense interest in the functional role that lipids might play in the plasma membrane. This project deploys a new class of synthetic lipid bilayers to begin drawing connections between the structure of lipid molecules-both in terms of individual molecular structure and supermolecular organization-and the function of the plasma membrane. The research tools developed and deployed here, called asymmetric giant unilamellar vesicles (AGUVs), are designed to uniquely mimic the cell membrane, capturing properties such as compositional asymmetry and molecular crowding better than other existing synthetic lipid bilayers. One of the many questions that AGUVs can help answer involves passive transport across the cell membrane. Passive transport is an important route for drug delivery and passage of environmental toxins into cells. Recent results show that the mechanism of this transport is complex, and highly dependent on lipid behavior. This project deploys an AGUV-based technique for systematically measuring the dynamics of solute molecules interacting with and penetrating lipid bilayers, yielding richer mechanistic data than other approaches are capable of delivering. AGUVs can also be used to study the mechanical properties of the cell membrane. These properties- particularly resistance to bending-control protein function and are important in a range of physiological processes. While synthetic lipid bilayers have been used to probe these properties, little is known about the effects of bilayer asymmetry on them. This project uses AGUVs to discover these effects. Lipid interactions with integral membrane proteins are likely a major mode by which lipids influence cell behavior. Very little is known, however, about the origins or controlling parameters of these interactions. This project begins to untangle this problem in AGUVs by using fluorescence microscopy to probe how peptides modeling the transmembrane regions of various proteins associate with segregated lipid domains. Finally, AGUVs can facilitate the systematic study of the effects of macromolecular crowding in the cell interior. The microfluidic technique by which AGUVs are formed allows for the inclusion of arbitrary molecules within them, leading to unique molecularly crowded structures.

描述（由申请人提供）：在过去的半个世纪中，分子生物学和生物化学的发展加深了我们对疾病的理解，并且这种理解推动了对生物技术和分子医学治疗方法的探索，这是解释分子生物学的主要理论支架。行为是结构与功能的关系。然而，我们对分子生物学结构与功能关系的理解在很大程度上仅限于中心法则的分子：蛋白质和寡核苷酸。中心法则之外的各种分子结构，特别是脂质和碳水化合物，表明这些也可以从结构驱动功能的角度来理解。特别是，人们对脂质在质膜中可能发挥的功能作用产生了浓厚的兴趣。该项目部署了一类新型的合成脂质双层，开始在脂质分子结构（无论是单个分子结构还是超分子组织）与质膜的功能之间建立联系。这里开发和部署的研究工具被称为不对称巨型单层囊泡（AGUV），旨在独特地模拟细胞膜，比其他现有的合成脂质双层更好地捕获成分不对称和分子拥挤等特性。 AGUV 可以帮助解决的众多问题之一涉及跨细胞膜的被动运输。被动转运是药物输送和环境毒素进入细胞的重要途径。最近的结果表明，这种运输的机制很复杂，并且高度依赖于脂质行为。该项目部署了一种基于 AGUV 的技术，用于系统测量溶质分子与脂质双层相互作用和穿透脂质双层的动力学，从而产生比其他方法能够提供的更丰富的机械数据。 AGUV 还可用于研究细胞膜的机械特性。这些特性（尤其是抗弯曲性）控制蛋白质功能，并且在一系列生理过程中很重要。虽然合成的脂质双层已被用来探测这些特性，但人们对双层不对称性对其的影响知之甚少。该项目使用 AGUV 来发现这些效应。脂质与完整膜蛋白的相互作用可能是脂质影响细胞行为的主要模式。然而，人们对这些相互作用的起源或控制参数知之甚少。该项目开始通过使用荧光显微镜来探究模拟各种蛋白质跨膜区域的肽如何与分离的脂质结构域相关联，从而解决 AGUV 中的这个问题。最后，AGUV 可以促进细胞内部大分子拥挤效应的系统研究。 AGUV 形成的微流体技术允许在其中包含任意分子，从而形成独特的分子拥挤结构。