STRUCTURE/FUNCTION STUDIES OF RETINOL BINDING PROTEINS

视黄醇结合蛋白的结构/功能研究

• 批准号: 2458863
• 负责人: MARCIA E. NEWCOMER
• 金额: $ 17.36万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-08-01 至 1999-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2458863
• 关键词: X ray crystallography; acyltransferase; chemical binding; computer simulation; conformation; esterification; hydropathy; intermolecular interaction; laboratory rat; ligands; microsomes; molecular polarity; molecular site; mutant; protein isoforms; protein structure function; proteolysis; retinaldehyde; retinoid binding proteins; retinoids; site directed mutagenesis; stop flow technique; vitamin metabolism; water

Vitamin A is required for a variety of biological processes. The studies described within the body of the grant proposal are directed at understanding how cellular retinol binding proteins function in retinol uptake, metabolism, and transport. It is our hypothesis that these proteins are more than just passive reservoirs which prevent non-specific partitioning of the hydrophobic vitamin into cellular membranes: The proteins themselves are involved in protein-protein interactions with the enzymes which metabolize retinoids, and as a consequence regulate retinol metabolism and function. With site-directed mutagenesis and a variety of techniques we will examine the following aspects of cellular retinol binding protein (CRBP) and cellular retinol binding protein type II(CRBP(II)): (1) How the intracellular transport proteins for vitamin A deliver retinol to the metabolizing enzymes. This is a process which requires protein-protein recognition. We will determine the sites on the carrier proteins involved in this recognition. (2) How the ligand enters the binding cavities of CRBP and CRBP(II). The X-ray structures are available for both proteins and it is apparent that a conformational change must accompany ligand entry and exit. We describe experiments aimed at determining the nature of this conformational change. (3) CRBP discriminates between retinol and retinal, whereas CRBP(II) does not. We will determine the structural basis for this functional difference. (4) The role of water in ligand binding. The binding cavities of CRBP and CRBP(II) contain specifically bound water molecules in addition to the ligand retinol. We postulate that there is a functional purpose, in addition to the obvious structural purpose, for the inclusion of water molecules in the binding site.

维生素A是多种生物过程所必需的。研究 在补助金提案的主体中描述的是针对 了解细胞视黄醇结合蛋白在视黄醇中的作用 吸收、代谢和运输。我们的假设是 蛋白质不仅仅是被动的储存库， 疏水性维生素进入细胞膜的分配： 蛋白质本身参与蛋白质-蛋白质相互作用， 代谢类维生素A并因此调节视黄醇的酶 代谢和功能。通过定点突变和各种 技术我们将研究细胞视黄醇的以下方面 结合蛋白和细胞视黄醇结合蛋白 II（CRBP（II））：（1）维生素A的细胞内转运蛋白 将视黄醇传递给代谢酶。这是一个 需要蛋白质-蛋白质识别我们将确定网站上的 参与这种识别的载体蛋白。(2)配体如何进入 CRBP和CRBP的结合腔（II）。X射线结构是 这两种蛋白质都可以得到，很明显， 变化必须伴随配体的进入和退出。我们描述的实验旨在 来确定这种构象变化的本质。(3)CRBP CRBP（II）区分视黄醇和视黄醛，而CRBP（II）不区分。我们 将决定这种功能差异的结构基础。（四） 水在配体结合中的作用。CRBP的结合腔和 CRBP（II）除了含有CRBP（II）外，还含有特异性结合的水分子。 配体视黄醇。我们假设有一个功能性的目的， 除了明显的结构目的外， 分子在结合位点。