LIPOPROTEIN STRUCTURE AND APOPROTEIN CONFORMATION

我们的长期目标是在分子细节上阐明血浆脂蛋白和载脂蛋白的结构、稳定性和动态特性，这对理解脂质相互作用、载脂蛋白交换、脂蛋白细胞表面相互作用、受体介导的脂蛋白摄取和脂蛋白相互转化至关重要。作为酶的辅助因子、受体的配体或胆固醇逆向转运的介质的稳定性及其功能作用。详细了解载脂蛋白结构的稳定性和适应性对于进一步了解脂蛋白的结构和功能至关重要。这种独特的结构适应性的精确分子机制尚不清楚，这将是拟议研究的重点。继续我们之前的工作，我们将专注于脂蛋白（HDL和LDL）和载脂蛋白的结构研究，以尽可能高的分辨率，使用最先进的分子生物物理学和结构生物学方法。将确定模拟可交换载脂蛋白（主要是apoA-I）序列中重要结构和功能单元的可交换肽序列中重要结构和功能单元的合成表达肽的结构和稳定相互作用。这些肽将基于包含天然载脂蛋白i的11/22残基螺旋片段，以及可交换载脂蛋白序列中基本11/22 mer串联重复序列的“理想”共识序列来设计。包含点和缺失突变体的突变形式的结构和动力学研究将集中在apoA-I分子的特定区域在其构象和稳定性中的作用。完整LDL的三维结构，重点是脂蛋白表面apo-B100的拓扑结构和分子构象，将通过冷冻电子显微镜和3d图像重建来确定。重点将是分析载脂蛋白b的组织和LDL颗粒的结构和功能域的定位，使用位点特异性免疫纳米金标记和结合LDL受体的直接可视化相结合。

Our long-term objectives are to elucidate in molecular detail the structure, stability and dynamic properties of the plasma lipoproteins and apolipoproteins that are vital to understanding of lipid interactions, apoprotein exchange, lipoprotein cell surface interactions, receptor- mediated lipoprotein uptake, and lipoprotein inter-conversions. The stabilization and their functional roles as cofactors for enzymes, ligands for receptors, or mediators of reverse cholesterol transport. A detailed understanding of apoprotein structural stability and adaptability is vital to further progress in understanding lipoprotein structure and function. The precise molecular mechanisms of this unique structural adaptability remain unclear, and will be the focus of the proposed research. Continuing our previous work, we will focus on structural investigations of lipoproteins (HDL and LDL) and apolipoproteins to highest possible resolution, using state-of-the-art methods of molecular biophysics and structural biology. The structure and stabilizing interactions of synthetic of expressed peptides that model important structural and functional units in the sequences of the exchangeable peptides that model important structural and functional units in the sequences of the exchangeable apoproteins (primarily apoA-I) will be determined. These peptides will be designed on the basis of the 11/22 residue helical segments comprising native apoA-I, and on an "idealized" consensus sequence for the fundamental 11/22-mer tandem repeat in the sequence of the exchangeable apoproteins. Structural and themodynamic studies of mutant forms of apoA-I encompassing point and deletion mutants will concentrate on the role of specific regions of the apoA-I molecule in its conformation and stability. The three-dimensional structure of intact LDL, with emphasis on the topology and the molecular conformation of the apo-B100 at the lipoprotein surface will be determined by cryo- electron microscopy and 3D-image reconstruction. The focus will be on the analysis of the organization of apo-B and the localization of structural and functional domains of the LDL particle, using a combination of site- specific immuno-nanogold labeling and direct visualization of the bound LDL receptor.