ISOPRENOID BIOSYNTHESIS IN THE RETINA

甲氧戊酸的异戊二烯途径产生了几个 对视网膜光感受器细胞功能至关重要的产品。这个 主要产品胆固醇约占组织的2%。 重量和约6-8%的总杆件外段(ROS) 膜脂。第二种产品，多氯乙酸磷酯(Dol-P)，是 N-连接糖蛋白中寡糖转移所必需的 合成，这是将视蛋白整合到 ROS膜。对这些物质的新陈代谢知之甚少 视网膜中的这两种脂类。本提案的目的是 缓解这一不足。胆固醇的化学水平， 高效毛细管电泳法测定角鲨烯、Dol-P和Dol-P 高效液相色谱。从头开始的角色 异戊二烯类化合物进入视网膜的合成和 ROS将通过测量以下各项的绝对速率确定 用~3H20和(~3H)乙酸乙酯在视网膜中合成。这个 ROS(A)组装对类异戊二烯合成的要求 膜，(B)视蛋白掺入ROS，和(C) 视蛋白的糖基化将使用抑制物进行检测。 在类异戊二烯从头途径的特定酶步骤 生物合成。最后，为了解决这个问题， 类异戊二烯和视蛋白由不同的途径传递， 功能性高尔基体衍生的囊泡交通依赖于 类异戊二烯向ROS的运输将被确定。 上述研究应该会产生大量的信息。 关于从头合成异戊二烯的作用和要求 ROS膜的组装和功能的合成。

The isoprenoid pathway from mevalonate generates several products important for retinal photoreceptor cell function. The primary product, cholesterol, represents about 2% of the tissue weight and about 6-8% of the total rod outer segment (ROS) membrane lipid. A second product, dolichyl phosphate (Dol-P), is required for oligosaccharide transfer in N-linked glycoprotein synthesis, a process essential for the integration of opsin into the ROS membrane. Very little is known regarding the metabolism of these two lipids in the retina. The present proposal is aimed at alleviating this deficiency. The chemical levels of cholesterol, squalene, Dol-P and dolichol will be determined by high performance liquid chromatography. The role of de novo synthesis in the incorporation of isoprenoids into the retina and ROS will be determined by measuring the absolute rates of synthesis in isolated retina using 3H20 and (3H)acetate. The requirement for isoprenoid synthesis for the (a) assembly of ROS membranes, (b) incorporation of opsin into ROS, and (c) glycosylation of opsin will be examined using inhibitors which act at specific enzymatic steps of the de novo pathway of isoprenoid biosynthesis. Finally, to address the question of whether isoprenoids and opsin are routed by different pathways, the dependence of functioning Golgi-derived vesicular traffic on the transport of isoprenoids to the ROS will be determined. The above studies should yield considerable information concerning the role and requirement of de novo isoprenoid synthesis for the assembly and function of ROS membranes.