Unraveling the Mechanism of Manganese (II) Oxidation by Pseudomonas Putida

揭开恶臭假单胞菌氧化锰 (II) 的机制

• 批准号: 0422232
• 负责人: Bradley Tebo
• 金额: $ 93万
• 依托单位: University of California-San Diego Scripps Inst of Oceanography
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-08-01 至 2006-05-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0422232&HistoricalAwards=false
• 关键词: Unraveling; Mechanism; Manganese; II; Oxidation

The oxidation of soluble manganese(II) to insoluble Mn(III) and Mn(IV) oxyhydroxides has a profound effect on the environment. The highly reactive Mn oxide mineral phases scavenge numerous heavy metals, oxidize various toxic organic and inorganic compounds, and serve as electron acceptors for growth of anaerobic bacteria. In nature, most Mn oxides form through the activities of microorganisms, primarily bacteria. Studies of three phylogenetically-diverse Mn(II)-oxidizing bacteria show that a homologous gene encoding a multicopper oxidase-like protein is required for Mn(II) oxidation. The direct link between those genes and enzymatic oxidation of Mn(II), however, has yet to be established. This project focuses on the mechanism, regulation and function of Mn(II) oxidation in Pseudomonas putida strain GB-1. Spectroscopic evidence indicates that Mn(II) oxidation proceeds via two sequential one-electron steps, both of which are enzymatically catalyzed. The recent discovery of a second multicopper oxidase gene required for Mn(II) oxidation in GB-1, in addition to the previously described cumA, suggests that two multicopper (Mn) oxidases drive this catalytic process. The new 5.9 Kb gene resembles the putative Mn oxidase gene, mnxG, from Bacillus SG-1 in its highly conserved copper binding sites. This project will determine whether each protein catalyzes one step of the two-electron oxidation, or alternatively, whether they work in concert to catalyze the overall reaction. Mutagenesis studies point to other factors that mediate Mn(II) oxidation, including interactions with c-type cytochromes, protein transport and location of the native enzymes. Physiological studies indicate that substrate-ligand binding and induction/ inhibition of oxidation by other metals, specifically Fe, play significant roles in Mn(II) oxidation. The results of this research will provide new insights into the reasons why bacteria oxidize manganese(II), broaden our understanding of biogeochemical cycles and the natural attenuation of toxic metals and organic compounds.Broader Impacts: The results from this project may lead to improved technologies for environmental remediation. Additionally, it will contribute to the education of undergraduate and highly-qualified high school students through independent research projects and mentorships; and to a training program for middle school teachers, highlighting the connections between the chemistry, biology and geology of the oceans.

可溶性锰（II）氧化为不溶性锰（III）和锰（IV）氧氢氧化物对环境有深远的影响。高活性的锰氧化物矿物相清除多种重金属，氧化各种有毒的有机和无机化合物，并作为厌氧细菌生长的电子受体。在自然界中，大多数锰氧化物是通过微生物（主要是细菌）的活动形成的。对三种系统发育不同的Mn（II）氧化细菌的研究表明，编码多铜氧化酶样蛋白的同源基因是Mn（II）氧化所必需的。然而，这些基因与酶促氧化Mn（II）之间的直接联系尚未确定。本项目主要研究恶臭假单胞菌菌株GB-1中Mn（II）氧化的机制、调控和功能。光谱证据表明，Mn（II）氧化通过两个连续的单电子步骤进行，这两个步骤都是酶催化的。除了先前描述的cumA外，最近还发现了GB-1中Mn（II）氧化所需的第二个多铜氧化酶基因，这表明两个多铜（Mn）氧化酶驱动了这一催化过程。这个5.9 Kb的新基因在高度保守的铜结合位点上与芽孢杆菌SG-1中推测的锰氧化酶基因mnxG相似。这个项目将确定每个蛋白质是否催化双电子氧化的一个步骤，或者它们是否协同催化整个反应。诱变研究指出了介导Mn（II）氧化的其他因素，包括与c型细胞色素的相互作用、蛋白质运输和天然酶的定位。生理研究表明，底物-配体结合和其他金属（特别是铁）的氧化诱导/抑制在Mn（II）氧化中起重要作用。本研究结果将为细菌氧化锰（II）的原因提供新的见解，拓宽我们对生物地球化学循环和有毒金属和有机化合物自然衰减的理解。更广泛的影响：该项目的结果可能会导致环境修复技术的改进。此外，它将通过独立的研究项目和指导为本科和高素质的高中学生的教育做出贡献；以及一个针对中学教师的培训项目，强调海洋的化学、生物学和地质学之间的联系。