CHARACTERIZATION OF MAMMALIAN FERROCHELATASE

本提案的目的是检查末端酶， 血红素生物合成途径、亚铁螯合酶（原血红素亚铁裂解酶，E.C. 4.99.1.1.），以确定有关的结构和催化信息 健康动物组织中的酶。 牛肝将是 酶，因为遗传原卟啉病已被描述在牛。 一旦 从正常动物中获得有关酶的信息，那么它将 可以检查来自患有以下疾病的动物的亚铁螯合酶 这样我们就可以了解它的分子基础 疾病 本文概述的实验方法涉及检查 铁螯合酶在膜中取向，可能的 蛋白质-脂质和蛋白质-蛋白质相互作用及其催化的研究 酶的功能。 数据将主要来自化学 纯化酶的修饰研究以及 在重构脂质体系统中的相互作用以及通过使用 卟啉底物类似物和抑制剂。 这个项目的最终目标 实验室是生化表征的三个终端膜 并将它们重组成脂质体， 它们在膜上是矢量组织的 在线粒体中。 该系统的研究首先在正常动物和 然后在斑岩动物中，沿着与临床医生的合作研究， 在这一领域，可能会导致生物化学和分子的理解， 原卟啉症和杂色卟啉症。

The objective of the present proposal is to examine the terminal enzyme in the heme biosynthetic pathway, ferrochelatase (protoheme ferrolyase, E.C. 4.99.1.1.), to determine structural and catalytic information about the enzyme from healthy animal tissue. Bovine liver will be the source of enzyme since a genetic protoporphyria has been described in cattle. Once information is obtained about the enzyme from normal animals, then it will be possible to examine ferrochelatase from animals suffering from protoporphyria so that we may then understand the molecular basis of that disease. The experimental approach outlined herein involves examination of the orientation of ferrochelatase in membranes, definition of possible protein-lipid and protein-protein interactions and study of the catalytic functioning of the enzyme. The data will come largely from chemical modification studies on the purified enzyme as well as examination of interactions in reconstituted liposomal systems and through the use of porphyrin substrate analogs and inhibitors. The ultimate goal of this laboratory is to biochemically characterize the three terminal membrane associated enzymes of this pathway and reconstitute them into liposomes so that they are vectorally organized across the membrane as they are in situ in the mitochondria. The study of the system first in normal animals and then in porphyric animals, along with collaborative studies with clinicians in he field, may lead to the biochemical and molecular understanding of protoporphyria and variegate porphyria.