Regulation of vitamin A metabolism in the eye

描述（由申请人提供）：维生素A是脊椎动物生物学中至少两种关键代谢物的前体，11-顺式视网膜，视觉发色基团和视黄酸，核受体的配体。膳食维生素A的吸收及其被目标身体组织吸收依赖于特殊的结合蛋白、转运蛋白和酶。参与代谢机制的关键成分是卵磷脂：视黄醇酰基转移酶（LRAT），一种催化视黄醇酯形成的酶。LRAT的生理作用引起了科学和临床的兴趣，因为这种酶是维持全身维生素A稳态和视觉发色团再生所必需的。LRAT也是开发组织特异性药物递送系统的有吸引力的靶点。然而，由于缺乏有关这些过程的分子基础的数据，在了解维生素A摄取和体内平衡的生物化学方面的进展受到阻碍。我们长期以来的研究兴趣集中在理解LRAT的催化和生理作用以及控制细胞类视黄醇摄取的机制。通过这一应用，我们拟阐明LRAT加工维生素A的基本分子机制。在Aim 1中，我们将描述区分LRAT与相关酶的分子适应性，以赋予其维生素A特异性。我们还将确定lrat特异性类视黄醇结合域和参与底物-蛋白质相互作用的关键残基。特异性目的2将侧重于确定LRAT酰基转移酶活性的分子机制。通过求解和分析HRASLS/LRAT嵌合蛋白的晶体结构，我们将发现导致酰基转移酶活性获得的关键分子适应。我们将在几个层面上定义这些变化，包括酶的一般拓扑结构，磷脂结合模式的调整，以及酶活性位点的水排斥机制。最后，在特定的Aim 3中，我们将检查酶的类视黄醇特异性。我们将分析嵌合酶在类维生素A结合状态下的结构，以获得有关维生素A结合位点的确切位置和组织的详细信息。总之，这些研究将提供新颖而全面的知识，填补我们对维生素A代谢和视觉发色团产生的理解空白。解剖类视黄醇酯化的分子机制将为更有效地治疗人类视网膜退行性疾病铺平道路。

DESCRIPTION (provided by applicant): Vitamin A is the precursor of at least two critical metabolites in vertebrate biology, 11-cis-retinal, the visual chromophore and retinoic acid, a ligand for nuclear receptors. Absorption of dietary vitamin A and its uptake by targeted body tissues depends on specialized binding proteins, transporters, and enzymes. Key components of the involved metabolic machinery are lecithin: retinol acyltransferase (LRAT), an enzyme that catalyzes the formation of retinyl esters. The physiological role of LRAT has attracted scientific and clinical interest because this enzyme is essential for maintaining systemic vitamin A homeostasis and regenerating visual chromophore. LRAT is also attractive targets for the development of a tissue-specific drug delivery system. Yet, progress in understanding the biochemistry of vitamin A uptake and homeostasis is hindered by a shortage of data regarding the molecular basis of these processes. We have a longstanding research interest focused on understanding LRAT's catalytic and physiological action as well as a mechanism that governs cellular retinoid uptake. Through this application, we propose to elucidate the fundamental molecular mechanism of vitamin A processing by LRAT. In Aim 1, we will delineate the molecular adaptations that discriminate LRAT from related enzymes to confer its vitamin A specificity. We also will identify the LRAT-specific retinoid binding domain and key residues involved in the substrate-protein interaction. Specific Aim 2 will focus on determining the molecular mechanism for LRAT's acyltransferase activity. By solving and analyzing crystal structures of HRASLS/LRAT chimeric proteins, we will discover the critical molecular adaptations that led to the acquisition of acyltransferase activity. We will define these changes at several levels, including the general topology of the enzyme, adjustments of the phospholipid binding mode, and the mechanism of water exclusion from the enzyme's active site. Finally, in specific Aim 3, we will examine the retinoid specificity of the enzyme. We will analyze the structures of the chimeric enzymes in their retinoid-bound states to obtain detailed information regarding the exact location and organization of the vitamin A binding site. Together, these studies will contribute novel and comprehensive knowledge, filling the gap in our understanding of the vitamin A metabolism and production of the visual chromophore. Dissecting molecular mechanisms underlying retinoid esterification will pave the way for more efficient treatment of human retinal degenerative diseases.