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

• 批准号: 8152258
• 负责人: NOAH MALMSTADT
• 金额: $ 29.25万
• 依托单位: UNIVERSITY OF SOUTHERN CALIFORNIA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-30 至 2015-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8152258
• 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; 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

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. PUBLIC HEALTH RELEVANCE: Cells are surrounded by membranes that consist of two layers of lipid molecules, and the chemical composition is different in these two layers. In this project, a new type of artificial cell membrane that is uniquely capable of mimicking this asymmetry is deployed to study a range of important biological processes, including drug transport into cells, mechanical deformation of the cell membrane, and interactions between lipid molecules and receptor proteins involved in cancer treatment and diabetes.

描述（由申请人提供）：在过去的半个世纪中，随着分子生物学和生物化学的发展，我们对疾病的理解不断加深，并且这种理解推动了生物技术和分子医学治疗方法的研究，解释分子行为的主要理论框架一直是结构-功能关系。然而，我们对分子生物学结构-功能关系的理解很大程度上局限于中心法则的分子：蛋白质和寡核苷酸。 中心法则之外的分子结构的多样性，特别是在脂类和碳水化合物中，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。本计画利用一种新的合成脂质双层，开始绘制脂质分子结构与质膜功能之间的关联，包括个别分子结构与超分子组织。在这里开发和部署的研究工具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可以帮助回答的许多问题之一涉及跨细胞膜的被动运输。被动转运是药物递送和环境毒素进入细胞的重要途径。最近的研究结果表明，这种转运的机制是复杂的，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，用于系统地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，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性能。这些特性-特别是抗弯曲-控制蛋白质功能，并且在一系列生理过程中是重要的。虽然合成的脂质双层已被用来探测这些属性，很少有人知道双层不对称对他们的影响。该项目使用AGUV来发现这些影响。 脂质与膜蛋白的相互作用可能是脂质影响细胞行为的主要方式。然而，我们对这些相互作用的起源或控制参数知之甚少。该项目开始解开这个问题，在AGUV使用荧光显微镜，以探测如何肽模拟各种蛋白质的跨膜区域与分离的脂质结构域。 最后，AGUV可以促进细胞内部大分子拥挤效应的系统研究。形成AGUV的微流体技术允许在其中包含任意分子，从而导致独特的分子拥挤结构。 公共卫生相关性：细胞被膜包围，膜由两层脂质分子组成，这两层的化学成分不同。在这个项目中，一种新型的人工细胞膜，能够独特地模拟这种不对称性，被用于研究一系列重要的生物过程，包括药物转运到细胞中，细胞膜的机械变形，以及脂质分子和参与癌症治疗和糖尿病的受体蛋白之间的相互作用。