STRUCTURAL STUDIES OF LIPID/ PROTEIN SYSTEMS

脂质/蛋白质系统的结构研究

• 批准号: 2749737
• 负责人: LEONARD J. BANASZAK
• 金额: $ 38.15万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-05-01 至 1999-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2749737
• 关键词: Agnatha; X ray crystallography; apolipoproteins; computer simulation; conformation; crystallization; egg /ovum; fatty acid binding protein; high density lipoproteins; lipid transport; mutant; nonblood lipoprotein; physical model; protein purification; protein reconstitution; protein structure function; retinoid binding proteins; site directed mutagenesis; steroid hormone receptor; structural biology; thermodynamics; vitamin receptor; vitellin

X-ray crystallography will be used to study the molecular structure of lipid:protein complexes ranging from extracellular proteins such as lipovitellin and apo Al to small intracellular lipid binding proteins. All the studies play a role in understanding the stabilization of lipid in an aqueous environment along with associated health related problems which involve the transient flux of lipids. Lipovitellin from lamprey oocytes is one of the crystalline proteins. It contains 15% w/w of lipid. The present model, though incomplete, has been obtained by x-ray and neutron diffraction data. The current focus is the assignment of the amino acid sequence to the electron density map. A second study will involve crystallographic studies of both apo Al, a constituent of HDL, and reconstituted apo Al :lipid particles. Using an existing form of recombinant proapo Al, a new expression system will eliminate the "pro" piece and add five histidine residues to the carboxy-terminal. This will permit a simple and large scale purification protocol. Another aspect of the studies focusses on a family of intracellular lipid binding proteins which are responsible for the translocation of fatty acids and retinoids within the cell. The background studies have shown that the hydrophobic compound is contained within an internalized water filled cavity.. Site- directed mutagenesis and crystallography will be used to gate ligand binding and to alter the internalized structured water. A protein which binds retinoic acid in a manner analogous to the fatty acid and retinol binding proteins will be analyzed by x-ray crystallography. The fourth and last study involves the purification and crystallization of the ligand binding domain of the nuclear receptor protein, RXR. The receptors for vitamin A analogues (RXR and RAR) play a dominant role in the development of a body plan as shown by teratogenic effects in embryos. These hormonal properties are the result of effects on the transcriptional rates of special targeted genes. Although structural studies on the segment of the protein which binds to DNA have been completed, no three dimensional data on the retinoic acid-binding domain is available. Studies of RXR are closely related to those of the intra- cellular retinoic acid binding protein mentioned above.

X射线结晶学将被用来研究分子结构 脂类：来自细胞外蛋白的蛋白质复合体，如 卵黄素和载脂蛋白结合到细胞内的小分子脂结合蛋白。 这些研究对了解脂质的稳定性起到了一定的作用。 在水环境中以及相关的健康相关问题 这涉及到脂类的瞬时流动。七鳃鳗卵黄蛋白 卵母细胞是一种结晶蛋白。它包含15%的w/w 脂类。目前的模型虽然不完整，但已通过x射线获得。 和中子衍射数据。当前的焦点是分配 氨基酸序列到电子密度图。第二项研究将 包括对高密度脂蛋白的成分载脂蛋白Al的结晶学研究， 重组载脂蛋白A1：脂质颗粒。使用现有形式的 重组Proapo Al，一种新的表达系统将消除“PRO” 在羧基末端插入5个组氨酸残基。这将是 允许一种简单和大规模的纯化方案。的另一个方面 研究的重点是细胞内脂结合蛋白家族 它们负责脂肪酸和维甲酸的移位 在细胞内。背景研究表明，疏水性 化合物包含在一个内化的充满水的空腔中。站点- 定向诱变和结晶学将用于配基的选择 结合并改变内化的结构水。一种蛋白质，它可以 以类似于脂肪酸和视黄醇的方式结合维甲酸 结合蛋白将通过X射线结晶学进行分析。第四次 而最后一项研究涉及到的提纯和结晶 核受体蛋白RXR的配基结合域。这个 维生素A类似物的受体(RXR和RAR)在 人体计划的发展，如在 胚胎。这些荷尔蒙特性是由于 特殊靶向基因的转录速率。虽然是结构性的 对与DNA结合的蛋白质片段的研究已经完成 完成后，没有关于维甲酸结合结构域的三维数据 是可用的。RXR的研究与Intra的研究密切相关 上面提到的细胞维甲酸结合蛋白。