Intrinsic curvature induced packing heterogeneity and non-uniform distribution of cholesterol and Abeta peptide in lipid bilayers

固有曲率诱导脂质双层中胆固醇和 Abeta 肽的堆积异质性和不均匀分布

• 批准号: 10656172
• 负责人: Radha Ranganathan
• 金额: $ 10.88万
• 依托单位: CALIFORNIA STATE UNIVERSITY NORTHRIDGE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-01 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10656172
• 关键词: Amyloid beta-Protein; Area; Behavior; Binding; Biological; Brain; Calibration; Characteristics; Charge; Cholesterol; Detection; Development; Electron Spin Resonance Spectroscopy; Electrostatics; Evolution; Exhibits; Fluorescence; Gel; Glycerol; Goals; Growth; Heterogeneity; Hydrophobicity; Individual; Knowledge; Length; Lipid Bilayers; Lipids; Liquid substance; Measures; Membrane; Membrane Fluidity; Methodology; Methods; Microscopy; Movement; Peptides; Pharmaceutical Preparations; Phase; Phosphorylcholine; Positioning Attribute; Property; Reporting; Research; Resolution; Shapes; Stress; Structure; Variant; Vesicle; experimental study; fluorophore; improved; innovation; laurdan; melting; neurotoxic; neurotoxicity; novel; preference; redshift; second harmonic

Project Summary: Lipid intrinsic curvature is a fundamental physical feature that regulates membrane interactions. The goal is to elucidate intrinsic curvature induced effects on the growth of membrane curvature and packing heterogeneity upon mixing lipids of dissimilar curvature and the consequent distribution of cholesterol and the neurotoxic 25-35 segment of Aβ peptides in bilayers, using a novel steady state fluorescence method that utilizes the computed second harmonic (SH) of the zeroth order spectra to better resolve spectral lines that are not obvious in the zeroth harmonic. The polarity sensitive fluorophore Laurdan will be employed. Laurdan emission in bilayers is a composite of a line, referred to as blue, from the denser packed region at 432 nm and a red line from the less packed region starting at 440 nm in single component gel phases evolving to 490 nm upon melting or mixing with a higher curvature lipid. This path of structure evolution was a new finding. The present hypothesis is: Any added molecule of a dissimilar curvature enters the less packed region of the bilayer. A higher curvature molecule increases stress which is then relieved by growth in flat domains. Laurdan’s preference for flat regions together with growth in such domains contribute to an increase in Laurdan presence in flat domains. Fluorescence spectra register an increase in the area, Ab, of the blue line from flat domains and decrease in the red line area, Ar, along with a red-shift of the red peak and no shift in the blue peak. A lower curvature molecule decreases stress and the membrane tends toward a homogeneous curvature distribution resulting in an Ab decrease, Ar increase, blue shift of the red peak and no shift of the blue peak. Enhancement of heterogeneity in regional curvature due to dissimilarity of intrinsic curvature of mixing lipids and preferential distribution of cholesterol or Aβ peptide that manifest distinct difference in behaviors of areas and peak positions are predicted. Laurdan fluorescence spectra will be measured without and with cholesterol or Aβ (25-35) in mixed bilayers of (i and ii) 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) with (i) 1-palmitoyl-2-oleoyl-sn- glycero-3-phosphocholine (POPC), and (ii) 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-(1′-rac-glycerol (POPG) and (iii) brain lipids. SH of spectra will be computed to determine the number of elementary emissions. Spectra will be fitted to this number of log-normal functions according to developed methodology to resolve spectral lines. Area and peak position changes with mixture composition will be examined to determine packing heterogeneity in the bilayers without the transmembrane molecules and the type of distribution (uniform or non-uniform) of these molecules. Computing the SH of fluorescence spectra by modulating the wavenumber in analogy to field modulation in ESR spectroscopy is an innovative approach to bring superior definition to features missed in broad featureless in measured zeroth harmonics. The proposed research will significantly enhance current knowledge by elucidating the key role of intrinsic curvature in the mechanism by which the added molecule enters, accumulates preferentially, inhomogenously rigidifies the bilayer and changes its shape.

项目摘要：脂质固有曲率是调节膜的基本物理特征 目的是阐明内在曲率对膜曲率增长的影响。 混合不同曲率的脂质时的堆积异质性以及随之而来的分布 使用新型稳态荧光检测双层中的胆固醇和 Aβ 肽的神经毒性 25-35 片段 利用计算出的零阶频谱的二次谐波 (SH) 来更好地解析频谱的方法 将使用极性敏感荧光团 Laurdan。 第 432 章 nm 和一条红线，来自填充较少的区域，从单组分凝胶相的 440 nm 开始，逐渐演变成 熔化或与更高曲率脂质混合后的波长为 490 nm，这种结构演化路径是一个新发现。 目前的假设是：任何添加的具有不同曲率的分子都会进入该区域的填充较少的区域。 较高曲率的分子会增加应力，然后通过劳丹区域的生长来缓解。 对平坦区域的偏好以及这些领域的增长有助于劳丹存在的增加 在平坦域中，荧光光谱显示平坦域中蓝线的面积 Ab 有所增加，并且 红线区域 Ar 减少，同时红色峰红移，而蓝色峰 A 没有变化。 曲率分子降低应力，膜趋于均匀曲率分布 导致 Ab 减少，Ar 增加，红色峰蓝移，而蓝色峰没有增强。 由于混合脂质和优先的固有曲率的不同而导致区域曲率的异质性 胆固醇或 Aβ 肽的分布，表现出区域和峰值位置行为的明显差异 预测 Laurdan 荧光光谱将在没有和有胆固醇或 Aβ (25-35) 的情况下进行测量。 (i 和 ii) 1,2-二棕榈酰-sn-甘油-3-磷酸胆碱 (DPPC) 与 (i) 1-棕榈酰-2-油酰-sn- 的混合双层 甘油-3-磷酸胆碱 (POPC)，和 (ii) 1-棕榈酰-2-油酰-sn-甘油-3-磷酸-(1'-rac-甘油 (POPG) (iii) 计算光谱的 SH 以确定元素发射的数量。 将根据已开发的方法拟合此数量的对数正态函数以解析谱线。 将检查面积和峰位置随混合物组成的变化以确定堆积异质性 在没有跨膜分子的双层中以及分布的类型（均匀或不均匀） 通过模拟场调制波数来计算荧光光谱的 SH。 ESR 光谱调制是一种创新方法，可为在 所测量的零次谐波广泛无特征。拟议的研究将显着增强电流。 通过阐明内在曲率在添加分子的机制中的关键作用来获得知识 进入、优先积聚、不均匀地使双层变硬并改变其形状。