SFG Investigation of Peptide and Protein Orientations in Membranes

膜中肽和蛋白质方向的 SFG 研究

• 批准号: 7662349
• 负责人: ZHAN CHEN
• 金额: $ 29.22万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-08-01 至 2013-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7662349
• 关键词: Affect; Alamethicin; Antibiotic Resistance; Antibiotics; Asthma; Attenuated; Bacteria; Biological; Biological Process; Calibration; Cell membrane; Cells; Charge; Classification; Communicable Diseases; Crystallization; Crystallography; Detection; Development; Disease; Drug resistance; Entropy; Environment; Escherichia coli; Fluorescence; Frequencies; Future; GABA Receptor; GTP-Binding Proteins; Generations; Heart Diseases; Heart failure; Heterotrimeric GTP-Binding Proteins; Hypertension; In Situ; Investigation; Ion Channel Protein; Ion Transport; Knowledge; Lead; Lipid Bilayers; Lipids; Literature; MSI-78; Measurement; Measures; Medicine; Membrane; Membrane Proteins; Methodology; Methods; Micelles; Modeling; Molecular; Mutagenesis; Opiate Addiction; Optics; Parkinson Disease; Peptides; Peripheral; Play; Property; Proteins; Research; Role; Sampling; Schizophrenia; Signal Transduction; Solutions; Spectroscopy, Fourier Transform Infrared; Spectrum Analysis; Structure; Sum; Surface; Techniques; Testing; Transmembrane Domain; Vertebral column; Work; X-Ray Crystallography; addiction; antimicrobial; antimicrobial peptide; base; biological systems; design; gamma-Aminobutyric Acid; hypertensive heart disease; improved; innovation; insight; interfacial; magainin; pardaxin; peptide G; peptide structure; prevent; protein complex; protein function; protein structure; public health relevance; receptor; relating to nervous system; solid state nuclear magnetic resonance; synthetic peptide

DESCRIPTION (provided by applicant): This research will develop a systematic way to use combined sum frequency generation (SFG), double resonance SFG (DRSFG), attenuated total refection FTIR (ATR-FTIR), and four-wave mixing (FWM) studies on alpha-helical structures to characterize membrane peptide/protein orientation in a single lipid bilayer in situ. The methodology can deduce the absolute orientation and complicated orientation distribution for a membrane peptide or protein in a single lipid bilayer that closely resembles the real membrane environment. It can also study the effect of asymmetry in lipid bilayers on peptide orientations, and can characterize structural information of peptides with very small surface coverage. The orientations of various antimicrobial peptides in lipid bilayers obtained in this study can help to elucidate modes of actions of such peptides on membranes. Antibiotic resistance is one of the most pressing problems in medicine at present, and we believe that this research will impact the design and optimization of peptides for antimicrobial purposes. The methodology will also be applied to study orientations of subunits of G-proteins in various environments, lending unique insight into how receptors and G proteins are organized in membranes during signal transduction and providing fundamental insights into various diseases such as cardiac failure. The specific aims are: 1. SFG studies supplemented by ATR-FTIR and FWM research can provide unique orientational information of various membrane peptides in a single lipid bilayer. These studies will lead to the determination of more detailed orientation distribution of the peptides in the membrane environment. The result here will also provide a calibration base for the studies proposed in Specific Aim 2. 2. DRSFG will be used to investigate the peptides examined in Specific Aim 1 to show that DRSFG can greatly improve the sensitivity of normal SFG. Unique structural information of membrane peptides with a very low surface concentration (peptide-lipid molar ratio<1:5,000) can be characterized using DRSFG. 3. In addition to ?-helical peptides, ?-helical structures in proteins will also be investigated to demonstrate the feasibility of determining structural information of secondary structural domains of membrane proteins and the orientation of membrane proteins using SFG, supplemented by ATR-FTIR and FWM. The G?1?2 subunit of a trimeric G-protein will be used as a model in this research. PUBLIC HEALTH RELEVANCE In this research a combination of vibrational spectroscopic techniques can provide vital orientational information regarding membrane peptides and proteins, which is difficult to obtain otherwise. Such work enables in-depth understanding of membrane orientations of antimicrobial peptides and G-proteins, providing important information to develop cures for infectious diseases, heart disease, asthma, opioid addiction, and hypertension.

描述（由申请人提供）：本研究将开发一种系统的方法，使用组合和频发生（SFG）、双共振SFG（DRSFG）、衰减全反射FTIR（ATR-FTIR）和四波混频（FWM）研究α-螺旋结构，以原位表征单个脂质双层中的膜肽/蛋白质取向。该方法可以推导出的绝对取向和复杂的取向分布的膜肽或蛋白质在一个单一的脂质双层，非常类似的真实的膜环境。它还可以研究脂质双层中的不对称性对肽取向的影响，并且可以表征具有非常小的表面覆盖度的肽的结构信息。在这项研究中获得的脂质双层中的各种抗菌肽的取向可以帮助阐明这些肽对膜的作用模式。抗生素耐药性是目前医学上最紧迫的问题之一，我们相信这项研究将影响抗菌肽的设计和优化。该方法还将应用于研究G蛋白亚基在各种环境中的取向，从而对受体和G蛋白在信号转导过程中如何在膜中组织提供独特的见解，并为心力衰竭等各种疾病提供基本见解。具体目标是：1.由ATR-FTIR和FWM研究补充的SFG研究可以提供单个脂质双层中各种膜肽的独特取向信息。这些研究将导致更详细的肽在膜环境中的取向分布的测定。这里的结果也将为具体目标2中提出的研究提供校准基础。2. DRSFG将用于研究特定目标1中检查的肽，以表明DRSFG可以大大提高正常SFG的灵敏度。具有非常低的表面浓度（肽-脂质摩尔比<1：5，000）的膜肽的独特结构信息可以使用DRSFG来表征。3.此外，？螺旋肽，本研究亦将研究蛋白质的螺旋结构，以证明利用SFG，辅以ATR-FTIR和FWM，来确定膜蛋白的二级结构域的结构信息和膜蛋白的取向的可行性。G？一个？2亚基的三聚体G蛋白将被用作本研究的模型。公共卫生相关性在这项研究中，振动光谱技术的组合可以提供有关膜肽和蛋白质的重要方向信息，这是难以获得的。这些工作使人们能够深入了解抗菌肽和G蛋白的膜取向，为开发传染病、心脏病、哮喘、阿片类药物成瘾和高血压的治疗方法提供重要信息。