Molecular breeding of a toxic chemicals-degrading-bacterium

1.可溶性甲基单氧酶（sMMO）基因被克隆自甲基有效的细菌、甲基化酶M （链M）和序列。（1） sMMO结构基因（约6 kb）及其启动区域已被测序，并被确认为sMMO基因通过比较从M.中提取的N终端氨基酸序列中纯化的sMMO基因的互补DNA序列而被比较。（2）某些特征被澄清为后续;· sMMO中的应变M与DNA序列中的甲基溴三孢子体OB 3b和甲基可可囊泡浴有关。金属绑定站点在sMMO的活跃中心，G/C含量高，G/C含量高，在第三封信中，以及在DNA序列中确认绑定站点。等离子体pWM 3编码sMMO基因被引入大肠杆菌。在E.大肠杆菌吸附pWM 3中没有检测到三氯乙烯降解活性，而在大肠杆菌吸附pWM 3之间没有发现三氯乙烯降解活性，SDS-PAGE宿主。羟基酶，排序的组件之一，有alpha 2 beta 2伽马2个子单位，并在活动中心所需的含有离子，指标是那些子单位的精确组装和金属需要进行快速水解酶活性。

1. Soluble methane monooxygenase (sMMO) gene was cloned from a methane-utilizing bacterium, Methylocystis sp.M (strain M), and sequenced.(1) sMMO structural gene (about 6kb) and its promotor region were sequenced, and those were confirmed to be sMMO genes by comparing them with complemental DNA sequences derived from N-terminal amino acid sequences of purified sMMO in strain M.(2) Some characteristics were cleared as follows ;・ sMMO from strain M was related closely with those of methylosinus trichosporium OB3b and Methylococcus capsulatus Bath in DNA sequences and putative amino acid sequences.・ Metal-binding site in a active center of sMMO,high G/C content in the third letter of the used codons, and prothsetic groups binding site were confirmed in that DNA sequence.2. Plasmid pWM3 coding sMMO genes was introduced into E.coli. Trichloroethylene degradation activity was not detected in E.coli harboring pWM3, and no difference in lysates was observed between E.coli harboring pWM3 and host by SDS-PAGE.Hydroxylase, one of the components of sMMO,consisted with alpha 2 beta 2 gamma 2 subunits and requires ferric ion in the active center, indicating that the precise assembly of those subunits and a metal would be required to express hydroxylase activity.