PARACOCCUS DENITRIFICANS IRON TRANSPORT APPARATUS

本节目将重点介绍铁的生物有机化学。 脱氮副球藻对副菌素的加工，即 微生物的铁摄取装置。特别强调的是 放在铁副肌动蛋白受体的作用上。我们的研究 到目前为止，已经揭示了1)对乙酰肌酸铁的动力学 促进铁摄取与“出租车”机制一致 其中，铁副肌动蛋白被吸附到 微生物、铁的释放和延缓的配体 2)微生物具有80kD的外膜 铁饥饿诱导的结合L铁蛋白的蛋白质 其表观解离常数为0.7uM，这是我们拥有的一种蛋白质 基本提纯；3)这种蛋白质不会从 和4)光亲和标记物，铁 L-对羟基苯甲酸叠氮铁可用于标记对羟基苯甲酸铁 受体蛋白。 有了这些发现和我们拥有的技术 开发，我们将解决以下问题，问题 虽然不是独一无二的，但主要集中在 铁副肌动蛋白受体。1)底物的具体程度 铁副肌动蛋白受体？2)是延缓的副肌动蛋白铁 释放为完整的副肌动蛋白？3)铁是如何从 铁副肌动蛋白与受体结合后？4) 受体蛋白的哪些结构特征对 铁副肌动蛋白的加工，即什么是关键的结构 负责结合的蛋白质成分？ 铁副肌动蛋白受体蛋白在一定程度上与 与最近对利用这些表面的兴趣有关 表达蛋白在疫苗研制中的应用 机会主义细菌。不幸的是，人们对此知之甚少 这些铁载体受体的分离和性质 这一计划将至少部分缓解这一问题。

This program will focus on the bioorganic chemistry of ferric parabactin processing by Paracoccus denitrificans, i.e., the microorganism's iron uptake apparatus. Special emphasis will be placed on the role of the ferric parabactin receptor. Our studies to date have revealed that 1) the kinetics of ferric parabactin promoted iron uptake are consistent with a "Taxi Cab" mechanism in which ferric parabactin is adsorbed to the surface of the microorganism, the iron liberated and the deferrated ligand released; 2) the microorganism has an 80kD outer membrane protein inducible by iron starvation that binds ferric L-parabactin with an apparent Kd of 0.7uM, a protein which we have substantially purified; 3) this protein does not release iron from ferric parabactin by itself; and 4) the photoaffinity label, ferric L-parabactin azide can be utilized to label the ferric parabactin receptor protein. With these findings in hand and with the techniques we have developed, we will address the following questions, questions which although not exclusively, are largely focused on the role on the ferric parabactin receptor. 1) How substrate specific is the ferric parabactin receptor? 2) Is deferrated ferric parabactin released as intact parabactin? 3) How is iron liberated from ferric parabactin after it has been bound to the receptor? 4) What structural features of the receptor protein are critical to ferric parabactin processing, i.e., what are the key structural components of the protein responsible for binding? The relevance of the ferric parabactin receptor protein is in part associated with the a recent interest in utilizing these surface expressed proteins in the development of vaccines for opportunistic bacteria. Unfortunately, little is known about the isolation and properties of these siderophore receptors, a situation which this program will alleviate at least in part.