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: 7533281
• 负责人: ZHAN CHEN
• 金额: $ 29.29万
• 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/7533281
• 关键词: Affect; Alamethicin; Antibiotic Resistance; Antibiotics; Asthma; Attenuated; Bacteria; Biological; Biological Process; Calibration; Cell membrane; Cells; Charge; Classification; Communicable Diseases; Complex; Crystallization; Crystallography; Depth; Detection; Development; Disease; Drug resistance; Entropy; Environment; Escherichia coli; Fluorescence; Frequencies; Future; GABA Receptor; GTP-Binding Proteins; Generations; Heart Diseases; Heart failure; Helix (Snails); 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; Numbers; Opiate Addiction; Optics; Parkinson Disease; Peptides; Peripheral; Play; Property; Proteins; Public Health; Purpose; Rate; Research; Role; Sampling; Schizophrenia; Signal Transduction; Solutions; Spectroscopy, Fourier Transform Infrared; Spectrum Analysis; Structure; Sum; Surface; System; Techniques; Testing; Transmembrane Domain; Vertebral column; Work; X-Ray Crystallography; addiction; antimicrobial; antimicrobial peptide; base; design; ear helix; gamma-Aminobutyric Acid; hypertensive heart disease; improved; innovation; insight; interfacial; magainin; pardaxin; peptide G; peptide structure; prevent; protein function; protein structure; receptor; relating to nervous system; size; solid state; 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，使用组合和频率产生（SFG），双共振SFG (DRSFG)，衰减全反射FTIR （ATR-FTIR）和四波混合（FWM）研究α -螺旋结构来原位表征单个脂质双分子层中的膜肽/蛋白质取向。该方法可以推导出膜肽或膜蛋白在单个脂质双分子层中的绝对取向和复杂取向分布，与真实的膜环境非常接近。它还可以研究脂质双分子层的不对称性对肽取向的影响，并可以表征表面覆盖很小的肽的结构信息。本研究获得的各种抗菌肽在脂质双层中的取向可以帮助阐明这些肽在膜上的作用模式。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不同环境下G蛋白亚基的取向，为信号转导过程中受体和G蛋白如何在膜中组织提供独特的见解，并为各种疾病（如心力衰竭）提供基本见解。具体目标是：1。SFG研究与ATR-FTIR和FWM研究相结合，可以提供单个脂质双分子层中各种膜肽的独特取向信息。这些研究将有助于确定更详细的多肽在膜环境中的取向分布。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将用于研究特异性靶1中检测的肽，以表明DRSFG可以大大提高正常SFG的敏感性。低表面浓度（肽脂摩尔比< 1:50 000）的膜肽的独特结构信息可以用DRSFG表征。3. 除了？-螺旋肽，？还将研究蛋白质中的-螺旋结构，以证明利用SFG，辅以ATR-FTIR和FWM来确定膜蛋白二级结构域的结构信息和膜蛋白取向的可行性。G 1 ?三聚体g蛋白的2个亚基将作为本研究的模型。在这项研究中，振动光谱技术的结合可以提供关于膜肽和蛋白质的重要定向信息，这是很难获得的。这项工作使我们能够深入了解抗菌肽和g蛋白的膜取向，为开发治疗传染病、心脏病、哮喘、阿片类药物成瘾和高血压的方法提供重要信息。