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细胞膜的基本物理特征 互动。目的是阐明固有曲率对膜曲率生长的影响。 以及不同曲率的脂类混合后的堆积非均质性以及由此产生的分布 用一种新的稳态荧光法在双层中研究胆固醇和β多肽的神经毒性25-35片段 一种利用计算的零阶谱的二次谐波(SH)来更好地分辨光谱的方法 在第零次调和中不明显的线条。将使用极性敏感的荧光团Laurdan。 双分子层中的Laurdan发射是一条线的合成物，称为蓝色，来自密度较高的填充区，在432 440 nm处开始的单组分凝胶相中的红线，演变为 490 nm，熔化或与较高曲率的脂类混合。这种结构进化的路径是一个新发现。 目前的假设是：任何添加的曲率不同的分子都进入了分子堆积较少的区域 双层的。较高的曲率分子会增加应力，然后在平坦的磁区中生长来缓解应力。劳尔丹的 对平坦区域的偏好以及这些领域的增长有助于增加Laurdan的存在 在平面域中。荧光光谱记录了来自平坦结构域的蓝线面积AB的增加，以及 红线面积Ar减小，同时红峰红移，蓝峰无移位。一个更低的 曲率分子降低应力，膜趋于均匀的曲率分布 导致AB降低，Ar增加，红峰蓝移，蓝峰不移。增强功能 混合脂的固有曲率不同导致的区域曲率的不均质性和择优 胆固醇或β多肽在区域和峰位上表现出明显差异的分布 是可以预测到的。Laurdan荧光光谱将在没有和有胆固醇或Aβ(25-35)的情况下测量 (I和II)1，2-二棕榈酰基-2-甘油-3-磷胆碱(DPPC)与(I)-1-棕榈酰基-2-油酰基-Sn... 甘油-3-磷胆碱(POPC)和(Ii)1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-(1′-rac-glycerol 和(Iii)脑脂。将计算光谱的SH以确定元素发射的数目。光谱 将根据开发的方法来拟合这个数目的对数正态函数来解析谱线。 将检查随着混合物组成而发生的面积和峰位变化，以确定填料的异质性 在没有跨膜分子和分布类型(均匀或非均匀)的双层中 这些分子。类比场调制波数计算荧光光谱的SH ESR波谱中的调制是一种创新的方法，可以为 在测量的零次谐波中没有宽泛的特征。拟议中的研究将显著增强目前的 通过阐明本征曲率在添加分子的机制中的关键作用来获得知识 进入，优先堆积，不均匀地使双层变硬并改变其形状。