Novel Responses to Oxygen by Anaerobic Microorganisms

厌氧微生物对氧气的新反应

• 批准号: 7025621
• 负责人: Michael W. Adams
• 金额: $ 26.95万
• 依托单位: UNIVERSITY OF GEORGIA
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-03-01 至 2009-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7025621
• 关键词: NAD(P)H dehydrogenase; Raman spectrometry; X ray crystallography; active sites; aerobiosis; anaerobic bacteria; anaerobiosis; catalyst; circular magnetic dichroism; electron nuclear double resonance spectroscopy; electron spin resonance spectroscopy; enzyme activity; enzyme complex; enzyme mechanism; free radical oxygen; functional /structural genomics; hydrogen peroxide; infrared spectrometry; iron sulfur protein; microarray technology; oxidative stress; oxidoreductase; protein structure function; respiratory burst oxidase; stress proteins; superoxides

DESCRIPTION (provided by applicant): By definition, aerobic organisms, both prokaryotic and eukaryotic, require molecular oxygen for energy conservation. This is a mixed blessing, however, as extremely reactive oxygen derivatives are produced during normal metabolism that can damage all cellular components. These so-called reactive oxygen species (ROS) have been implicated in a wide variety of chronic and infectious human diseases, including cancer, Alzheimer's disease, arthritis and AIDS, yet ROS are also used as a defense system against pathogens and in signal transduction pathways. Understanding the responses of microbes to oxygen has direct ramifications for the treatment of diseases caused by anaerobic pathogens. In 1999 we proposed that anaerobes have a novel response to ROS in which a non-heme iron protein termed superoxide reductase (SOR) played a key role. SOR was characterized from the hyperthermophilic anaerobe, Pyrococcus furiosus, and over the prior funding period it has been established using structural and spectroscopic approaches that SOR is uniquely suited to catalyze superoxide reduction. Using DNA microarrays to all 2065 ORFs in the complete P. furiosus genome, it was shown that the genes encoding SOR and related proteins are all expressed at significant levels in the absence of any oxidative shock. P. furiosus is therefore continuously 'armed' and ready to deal with ROS exposure. This is a first line of defense, however, as DNA microarray analyses show that the complete response to oxidative stress requires the induction of a large number of novel proteins (encoded by conserved/hypothetical genes), some of which are also induced by growth at sub-optimal temperatures. In the proposed research, the novel stress-regulated proteins, together with SOR and related reductases and oxidases, will be characterized with respect to their regulation, multiprotein complex formation, and catalytic functions using immunological, biochemical and structural analyses. A variety of complementary spectroscopic techniques, including EPR, ENDOR, MCD, resonance Raman, FTIR and X-ray absorption, will be utilized to probe the catalytic function of specific members of the stress-related pathways, with particular emphasis on SOR. The results will provide completely new insights into the stress responses of anaerobes, and provide strategies for determining the function of uncharacterized hypothetic algenes that typically account for half of a microbial genome.

描述（由申请人提供）：根据定义，需氧生物（原核生物和真核生物）都需要分子氧来保存能量。然而，这是一个好坏参半的事情，因为在正常新陈代谢过程中会产生极活性的氧衍生物，可能会损害所有细胞成分。这些所谓的活性氧 (ROS) 与多种慢性和传染性人类疾病有关，包括癌症、阿尔茨海默病、关节炎和艾滋病，但 ROS 也被用作针对病原体的防御系统和信号转导途径。了解微生物对氧气的反应对于治疗厌氧病原体引起的疾病具有直接影响。 1999 年，我们提出厌氧菌对 ROS 有一种新的反应，其中一种称为超氧化物还原酶 (SOR) 的非血红素铁蛋白发挥了关键作用。 SOR 的特征来自于超嗜热厌氧菌，激烈火球菌，在之前的资助期间，已经使用结构和光谱方法确定了 SOR 独特地适合催化超氧化物还原。使用 DNA 微阵列分析完整的 P. Furiosus 基因组中的所有 2065 个 ORF，结果表明，在没有任何氧化休克的情况下，编码 SOR 和相关蛋白的基因均以显着水平表达。因此，P. Furiosus 不断“武装”并准备好应对 ROS 暴露。然而，这是第一道防线，因为 DNA 微阵列分析表明，对氧化应激的完全反应需要诱导大量新蛋白质（由保守/假设基因编码），其中一些蛋白质也是由生长诱导的次优温度。在拟议的研究中，将使用免疫学、生化和结构分析来表征新型应激调节蛋白以及 SOR 和相关还原酶和氧化酶的调节、多蛋白复合物形成和催化功能。各种互补的光谱技术，包括 EPR、ENDOR、MCD、共振拉曼、FTIR 和 X 射线吸收，将用于探测应激相关途径特定成员的催化功能，特别是 SOR。这些结果将为厌氧菌的应激反应提供全新的见解，并为确定通常占微生物基因组一半的未表征的假设藻类基因的功能提供策略。