Development and Characterization of Biotransformation System toward Helogenated Compounds with Yeast

(1)氯代乙烷和乙烯是一类特别有问题的污染物，生物处理方法越来越受到人们的关注。与三氯乙烯和五氯乙烷，无细胞膜组分的初始速率是一个数量级高于每单位酶与完整的酵母。六氯乙烷减少在可比率的膜和酵母。三氯乙烯被氧化成2，2，2-三氯乙醛和2，2，2-三氯乙醇。完整的酵母产生更多的三氯乙醇，动力学研究表明，三氯乙醛。五氯乙烷经还原脱卤生成三氯乙烯，三氯乙烯部分氧化生成三氯乙醛和三氯乙醇。六氯乙烷进行还原脱卤 ...更多信息 反应以产生四氯乙烯和五氯乙烷的混合物。(2)我们研究了表达大鼠肝细胞色素P450 1A 2（P450 1A 2）突变蛋白的重组酿酒酵母菌株是否有效地催化三氯乙烯、五氯乙烷和六氯乙烷的脱卤。结果发现，在一个假定的远端位点的突变P450 1A 2极大地提高营业额对这些底物无论是在有氧或厌氧条件下。例如，在推定的远端位点处的Thr 319 Ala突变在好氧条件下显著地提高三氯乙烯和五氯乙烷的降解速率，分别为2倍和2.7倍。Thr 319 Ala突变也强烈促进了对六氯乙烷的反应高达13倍和4.5倍，分别在有氧和厌氧条件下。结合反应机理讨论了Thr 319在这些氯化烃的催化作用中的作用。少

(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