Structure Function Relationship in Hemeproteins

血红素蛋白的结构功能关系

• 批准号: 6621301
• 负责人: Syun-Ru Yeh
• 金额: $ 27.63万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-12-01 至 2005-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6621301
• 关键词: Escherichia coli; Mycobacterium bovis; Mycobacterium tuberculosis; Raman spectrometry; bacterial proteins; enzyme activity; enzyme complex; enzyme mechanism; hemoglobin; ligands; microorganism metabolism; nitric oxide; oxygen consumption; oxygen microelectrode; prostaglandin endoperoxide synthase; protein protein interaction; protein structure function; site directed mutagenesis; ultraviolet spectrometry

The objective of this proposal is to elucidate the protein-ligand interactions and structure/function relationships in three new bacterial hemoglobins (Hb) and two mammalian prostaglandin H synthases (PGHS-1 and PGHS-2). The two bacterial hemoglobins from Mycobacterium tuberculosis (HbN and HbO) belong to a newly discovered truncated hemoglobin family, which are characterized by a novel two-over-two alpha-helical sandwich motif, the absence of the A- helix and the presence of an extended loop substituting for most of the F-helix. The physiological functions of HbN and HbO are not established but because O2 delivery in unicellular organisms is a diffusion-controlled process, functions other than oxygen transport have been put forth. The bacterial hemoglobin from E. coli (Hmp) is a flavohemoglobin consisting of a heme-containing globin-like domain and a FAD-containing reductase domain. It is believed that the function of Hmp is to detoxify NO and other reactive nitrogen species. The structural properties of the three bacterial hemoglobins will be fully characterized. Based on our preliminary resonance Raman studies, we postulate that the heme pockets of these bacterial hemoglobins are tailored to perform chemistry, such as oxygen activation, and that they may share structural and functional similarities to peroxidases. This hypothesis will be tested by studies of the reactions of these hemoglobins with NO, hydrogen peroxide and peroxynitrite. The possible role of these hemoglobins in protecting the microorganisms against attack by reactive nitrogen intermediates will be explored by monitoring NO and O2 consumption in wild-type and hemoglobin knock-out cells. Related reactions will be studied in the peroxidase sites of two PGHS isoforms, which play an essential role in the synthesis of prostaglandins. Preliminary data suggests that the proximal bond that coordinates the heme to the polypeptide is quite weak in PGHS's, which is very unusual for peroxidases. Experiments are proposed to test the functional consequences of this finding. These hemeprotein systems provide an excellent model for investigating fundamental structural properties that underlie biological reactivity.

该建议的目的是阐明三种新的细菌血红蛋白（Hb）和两个哺乳动物前列腺素H合酶（PGHS-1和PGHS-2）中的蛋白质 - 配体相互作用和结构/功能关系。 来自结核分枝杆菌（HBN和HBO）的两种细菌血红蛋白属于新发现的截短的血红蛋白家族，其特征是一种新型的两次两次α-甲基壳图主题，A-Helix的缺乏，是A-Helix的缺乏，并且在大部分for for For for for for for for for for for for for for for for for for for for forix themix botif。 HBN和HBO的生理功能不是建立的，而是因为单细胞生物中的O2递送是一个扩散控制的过程，因此提出了除氧运输以外的其他功能。 来自大肠杆菌（HMP）的细菌血红蛋白是由含血红素的球蛋白样结构域和含有FAD的还原酶结构域组成的黄素血红蛋白。 据认为，HMP的功能是对NO和其他反应性氮种类排毒。三种细菌血红蛋白的结构特性将得到充分表征。 根据我们的初步共振拉曼研究，我们假设这些细菌血红蛋白的血红素口袋是针对化学量的，例如氧气激活，并且它们可能与过氧化物酶具有结构和功能相似性。 该假设将通过研究这些血红蛋白与NO，过氧化氢和过氧亚硝酸盐的反应进行检验。这些血红蛋白在保护微生物免受反应性氮中间体攻击中的可能作用将通过监测野生型和血红蛋白敲除细胞中的NO和O2消耗来探索。相关反应将在两个PGHS同工型的过氧化物酶位点进行研究，这在Prostaglandins的合成中起着至关重要的作用。 初步数据表明，将血红素与多肽配位的近端键在PGHS中非常弱，这对于过氧化物酶来说是非常不寻常的。 提出了实验来测试该发现的功能后果。 这些血蛋白系统为研究生物反应性的基本结构特性提供了一个极好的模型。