Development and Characterization of Biotransformation System toward Helogenated Compounds with Yeast

酵母异构化化合物生物转化系统的开发和表征

基本信息

批准号：
07558083
负责人：
SHIMIZU Toru
金额：
$ 11.52万
依托单位：
Tohoku University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A)
财政年份：
1995
资助国家：
日本
起止时间：
1995 至 1996
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-07558083/
关键词：
Recombinant DNA Cytochrome Environment Bioremediation Bioreactor Halogenated Compounds

项目摘要

(1) Chlorinated ethanes and ethenes are a particularly problematic class of pollutants for which biological treatment methods are increasingly being sought.A recombinant Saccharomyces cerevisiae strain expressing rat liver cytochrome P450 1A2 oxidized tirchloroethylene and reduced pentachloroethane and hexachloroethane. With trichloroethylene and pentachloroethane, the initial rates of cell-free membrane fractions were an order of magnitude higher per unit enzyme than those obtained with intact yeast.Hexachloroethane was reduced at comparable rates by both membranes and yeast. Trichloroethylene was oxidized to 2,2,2-trichloroacetaldehyde and 2,2,2-trichloroethanol. Intact yeast produced proportionally more trichloroethanol which kinetic studies indicated arose from trichloroacetaldehyde. Pentachloroethane underwent ruductive dehalogenation to yield trichloroethylene, part of which was oxidized to trichloroacetaldehyde and trichloroethanol. Hexachloroethane underwent reductive dehalogen … More ation to yield a mixture of tetrachloroethylene and pentachloroethane.(2) We examined whether or not a recombinant Saccharomyces cerevisiae strain expressing rat liver cytochrome P450 1A2 (P450 1A2) mutant proteins efficiently catalyze dehalogenation of trichloroethylene, pentachloroethane, and hexachloroethane. It was found that mutations at a putative distal site of P450 1A2 enormously enhanced turnover numbers toward those substrates both under the aerobic or anaerobic conditions. For example, a Thr319Ala mutation at the putative distal site markedly enhanced the degradation rate of trichloroethylene and pentachloroethane by 2-and 2.7-fold.respectively, under the aerobic conditions. The Thr319Ala mutation also strongly facilitated the reaction toward hexachloroethane up to 13-and 4.5-fold both under the aerobic and anaerobic conditions, respectively. Role of Thr319 in the cataklytic functions toward those dhlorinated hydrocarbons are discussed in association with reaction meachanism. Less

（1）氯化乙烯和乙烯是一类特别有问题的污染物类别，生物治疗方法越来越成为扫描剂。一种重组的酿酒酵母表达大鼠肝细胞色素P450 1A2氧化的tirchorene和五氯乙烯和五氯乙烯乙烯和远射素。使用三氯乙烯和五氯乙烷，无细胞的膜级分的初始速率比完整酵母菌获得的每单位酶的数量级高。膜和酵母都以可比的速率降低了甲乙烷。将三氯乙烯氧化为2,2,2-三氯乙醛和2,2,2-三氯乙醇。完整的酵母产生了比例的三氯乙醇，动力学研究表明是由三氯乙醛引起的。五氯乙烷经过了固定的去解式化，产生三氯乙烯，其中一部分被氧化为三氯乙醛和三氯乙醇。 Hexachloroethane underwent reduced dehalogen… More ation to yield a mixture of tetrachloroethylene and pentachloroethane.(2) We examined whether or not a recombinant Saccharomyces cerevisiae strain expressing rat liver cytochrome P450 1A2 (P450 1A2) mutant proteins efficiently catalyze dehalogenation of trichloroethylene,五氯乙烷和六氯乙烷。已经发现，在有氧或厌氧条件下，P450 1A2的推定不同位点的突变增加了对这些底物的突变。例如，在推定的不同位点处的THR319ALA突变显着提高了三氯乙烯和五氯乙烷的降解速率，在有氧条件下，尊贵的态度。 THR319ALA突变还强烈支持对六氯乙烷的反应，分别在有氧和厌氧条件下分别为13倍和4.5倍。与反应机制有关，讨论了THR319在cataklytic函数中对那些脱氯的碳氢化合物的作用。较少的