FLUORESCENCE STUDIES OF BIOPHYSICAL PROPERTIES OF POLYUN

Polyun生物物理性质的荧光研究

• 批准号: 6160344
• 负责人: Burton J Litman
• 金额: --
• 依托单位: NATIONAL INSTITUTE ON ALCOHOL ABUSE AND ALCOHOLISM
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6160344
• 关键词: calorimetry; chemical models; ethanol; fluorescence; fluorescent dye /probe; lipid bilayer membrane; long chain fatty acid; membrane activity; membrane lipids; membrane structure; phase change; phosphatidylcholines; phospholipids; retina; synaptosomes; unsaturated fatty acids

Neuronal and retinal tissue are high in phospholipids containing either one or two polyunsaturated acyl chains. Our studies have led to the development of a novel model for phospholipid acyl chain packing, which involves the formation of dynamic lateral domains in the surface of the membrane. Differential scanning calorimetry (DSC) of mixtures of a disaturated (di16:0) phospha-tidylcholine (PC) and a dipolyunsaturated (di22:6n3) PC were studied. These lipids show poor miscibility in the liquid crystalline (LC) phase and demonstrate lateral phase separation in the gel phase. Fluorescence lifetime and anisotropy measurements, using the probe diphenylhexatriene (DPH), also indicates the presence of lateral heterogeneity in the membrane surface in both the gel and liquid crystal states. These studies support the presence of clusters or domains in mixtures of mixed chain and dipolyunsaturated phospholipids, as exist in both synaptosomal and retinal membranes. The propensity to form domains increases with increasing levels of acyl chain unsaturation, with 22:6n3 acyl showing the greatest level of domain formation. Both DSC and fluorescence data demonstrate that cholesterol preferentially interacts with saturated acyl chains and would, therefore, tend to concentrate in the more saturated regions of the membrane where it would effect both the size and lifetime of the domains. These findings have potentially important implications for integral membrane protein function. Domain formation would create regions of the membrane rich in highly unsaturated acyl chains. Proteins located in these regions would have different functional properties than those in more saturated regions of the membrane. Many neurotransmitter receptors and visual pigments are members of the G protein coupled-receptor family and reside in membranes containing very high levels of polyunsaturated acyl chains where domains of the type described are likely to occur. DSC measurements indicate that polyunsaturated phospholipids are more sensitive to the perturbing nature of ethanol. Therefore, the formation of highly unsaturated domains could result in regions of the synaptosomal membranes which are particularly sensitive to alcohols and other membrane soluble agents. Fluorescent probes have been widely used to characterize the acyl chain packing properties of lipid bilayers in both model and biological membranes. We have extended the analysis of the time-resolved anisotropy decay of one of the most commonly used probes, DPH, to allow a measure of the acyl chain packing order in the membrane interior vs the portion of the chain closer to the interfacial region. These data complement the information from NMR and characterize the time averaged properties of the ensemble of molecules making up the bilayer. This analysis allows a more refined discrimination of the packing properties of phospholipid acyl chains and their interaction with cholesterol.

神经细胞和视网膜组织中的磷脂含量很高，含有 一个或两个多不饱和酰基链。我们的研究导致了 一种新的磷脂酰链堆积模型的开发 涉及到在表面形成动态的侧向区域 薄膜。差示扫描量热法(DSC)测定有机硅混合物 饱和(di16：0)磷脂酰胆碱(PC)和二聚不饱和 对(di22：6N3)PC进行了研究。这些脂类显示出在 液晶(LC)相，并显示横向相分离 在凝胶阶段。荧光寿命和各向异性测量， 使用探针二苯基己三烯(DPH)，也表明存在 凝胶和液体中膜表面的横向非均质性 晶态。这些研究支持星系团的存在或 混合链和二多不饱和磷脂混合物中的结构域， 在突触体膜和视网膜膜中都存在。倾向于…… 形成结构域随着酰链不饱和度水平的增加而增加， 其中22：6n3酰基显示最大程度的结构域形成。两者都有 DSC和荧光数据表明，胆固醇优先 与饱和的酰基链相互作用，因此倾向于 浓缩在膜的较饱和区域，在那里它会 影响域的大小和寿命。这些发现有 完整膜蛋白潜在的重要意义 功能。结构域的形成将在膜上形成富含 高度不饱和的酰基链。位于这些区域的蛋白质会 具有与更饱和区域不同的功能特性 膜的表面。许多神经递质受体和视觉色素 G蛋白偶联受体家族的成员，存在于膜上 含有非常高水平的多不饱和酰基链，其中结构域 所描述的类型很可能会发生。差示扫描量热仪测量表明 多不饱和磷脂对扰动更敏感 乙醇的性质。因此，形成了高度不饱和的 结构域可以导致突触体膜的区域 对醇类和其他膜溶解剂特别敏感。 荧光探针已被广泛用于表征酰基链 脂类双层在模型和生物中的堆积性质 膜。我们推广了时间分辨各向异性的分析 最常用的探头之一DPH的衰变，以允许测量 膜内部的酰基链堆积顺序与部分 离界面区域更近的链条。这些数据补充了 来自核磁共振的信息，并表征了时间平均特性 组成双分子层的分子集合。这种分析允许更多的 磷脂酰基填充性的精细判别 链及其与胆固醇的相互作用。