Copper Bioavailability on Microbial Degradation of Trichloroethylene

铜对三氯乙烯微生物降解的生物利用度

• 批准号: 9708557
• 负责人: Jeremy Semrau
• 金额: $ 44万
• 依托单位: Regents of the University of Michigan - Ann Arbor
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-09-01 至 2002-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9708557&HistoricalAwards=false
• 关键词: Copper; Bioavailability; Microbial; Degradation; Trichloroethylene

9708557 Semrau This proposal was submitted in response to the Environmental Geochemistry and Biogeochemistry solicitation NSF 96-152, and is being funded jointly by the Divisions of Molecular and Cellular Biosciences, Chemistry and Earth Sciences, and by the MPS Office of Multidisciplinary Activities. This research will determine how soil surfaces limit the bioavailability of copper to methanotrophs and the resulting effect on the expression and activity of both forms of methane monooxygenase (MMO). Copper to biomass ratio is a key factor in regulating expression and activity of soluble and particulate methane monooxygenases (sMMO and pMMO, respectively) in liquid cultures of methanotrophs. To determine how different soil surfaces can limit the bioavailability of copper and the resulting effect on MMO expression and activity, two representative methanotrophs, M. trichosporium OB3b and M. albus BG8 will be grown in pure cultures in the presence of aluminum oxide (a representative of commonly occurring metal oxides) and montmorillonlte (representative of an important ion-exchange site bearing mineral) under a variety of solution conditions. These organisms were chosen for initial study because M trichosporium OB3b can express both forms of MMO while M. albus BG8 can express only pMMO. Experimental measurements include: speciation and amount of aqueous, adsorbed and cell-associated copper using atomic adsorption spectroscopy or a copper ion electrode; copper sorption by X-ray absorption spectroscopy; sMMO and pMMO expression by naphthalene oxidation assays and Western blots; propylene oxide assays of soluble and particulate fractions to localize and measure MMO activity; GC-MS analysis to determine the ability of these cells to oxidize methane and trichloroethylene. In subsequent experiments, artificial mixed cultures of both organisms will be grown in similar soil/solution conditions to understand how surfaces can cause population fluctuations by acting as a sink for copper. Experimental mea surements include: expression and activity of sMMO and pMMO; population composition by Western blot, acridine orange direct count, and PCR amplification of the genes for 16s rRNA, sMMO and pMMO; and the ability of mixed cultures to oxidize methane and trichloroethylene. This multidisciplinary research incorporates approaches from molecular biology, biochemistry and geochemistry, and is a model system for examining the effect of bioavailability of other metals on microbial degradation of both aromatic and aliphatic halogenated hydrocarbons. ***

9708557 塞姆劳 该提案是应NSF 96-152环境地球化学和生物地球化学招标而提交的，由分子和细胞生物科学、化学和地球科学部门以及MPS多学科活动办公室共同资助。 这项研究将确定土壤表面如何限制铜对甲烷氧化菌的生物利用度，以及对两种形式的甲烷单加氧酶（MMO）的表达和活性的影响。铜与生物量的比例是调节甲烷氧化菌液体培养物中可溶性和颗粒性甲烷单加氧酶（分别为sMMO和pMMO）表达和活性的关键因素。为了确定不同的土壤表面如何限制铜的生物有效性以及由此产生的对MMO表达和活性的影响，两种代表性的甲烷氧化菌，M。trichosporium OB 3b和M.在各种溶液条件下，在氧化铝（通常存在的金属氧化物的代表）和蒙脱石（具有重要离子交换位点的矿物的代表）存在下，在纯培养物中生长白血球BG 8。这些生物体被选择用于初始研究，因为发孢M. albus BG 8只表达pMMO。实验测量包括：使用原子吸收光谱法或铜离子电极测定含水、吸附和细胞相关铜的形态和量;通过X射线吸收光谱法测定铜吸附;通过萘氧化测定和蛋白质印迹法测定sMMO和pMMO表达;可溶性和颗粒级分的环氧丙烷测定以定位和测量MMO活性; GC-MS分析以确定这些细胞氧化甲烷和三氯乙烯的能力。在随后的实验中，这两种生物的人工混合培养物将在类似的土壤/溶液条件下生长，以了解表面如何通过充当铜的汇而引起种群波动。实验测量包括：sMMO和pMMO的表达和活性;通过Western印迹、吖啶橙子直接计数和PCR扩增16 s rRNA、sMMO和pMMO基因的群体组成;以及混合培养物氧化甲烷和三氯乙烯的能力。这一多学科的研究结合了分子生物学，生物化学和地球化学的方法，是一个模型系统，用于研究其他金属的生物利用度对芳香族和脂肪族卤代烃微生物降解的影响。 ***