从节杆菌菌株 J-1 中分离纯化苯甲腈酶及其性质研究。
Purification and Characterization of Benzonitrilases from Arthrobacter sp. Strain J-1.
机构信息
Department of Agricultural Chemistry and Research Center for Cell and Tissue Culture, Kyoto University, Sakyo-Ku, Kyoto 606, Japan.
出版信息
Appl Environ Microbiol. 1986 Feb;51(2):302-6. doi: 10.1128/aem.51.2.302-306.1986.
We found two kinds of benzonitrilases, designated benzonitrilases A and B, in a cell extract of Arthrobacter sp. strain J-1 grown on benzonitrile as a sole carbon and nitrogen source. Benzonitrilases A and B were purified approximately 409-fold and 38-fold, respectively. Purified benzonitrilase A appeared to be homogeneous according to the criteria of polyacrylamide gel electrophoresis. Both the enzymes hydrolyzed benzonitrile to benzoic acid and ammonia without forming benzamide as an intermediate. The molecular weights of benzonitrilases A and B were found to be 30,000 and 23,000, respectively. The subunit molecular weight of benzonitrilase A was the same as its molecular weight. The isoelectric points of benzonitrilases A and B were 4.95 and 4.80, respectively. The optimum temperature and pH, respectively, for benzonitrilase A were 40 degrees C and 8.5, and those for benzonitrilase B were 30 degrees C and 7.5. The K(m) values for benzonitrilases A and B were 6.7 mM and 4.5 mM, respectively. Both the enzymes degraded p-tolunitrile, 4-cyanopyridine, and p-chlorobenzonitrile, but they did not attack aliphatic nitriles or amides. Both the enzymes were inhibited by thiol reagents.
我们从以苯甲腈为唯一碳源和氮源的节杆菌（Arthrobacter sp.）J-1 细胞提取物中发现了两种苯甲腈酶，分别命名为苯甲腈酶 A 和 B。苯甲腈酶 A 和 B 分别被纯化了约 409 倍和 38 倍。根据聚丙烯酰胺凝胶电泳的标准，纯化的苯甲腈酶 A 似乎是均一的。两种酶都将苯甲腈水解生成苯甲酸和氨，而不形成中间产物苯甲酰胺。苯甲腈酶 A 和 B 的分子量分别为 30000 和 23000。苯甲腈酶 A 的亚基分子量与其分子量相同。苯甲腈酶 A 和 B 的等电点分别为 4.95 和 4.80。苯甲腈酶 A 的最适温度和 pH 值分别为 40°C 和 8.5，苯甲腈酶 B 的最适温度和 pH 值分别为 30°C 和 7.5。苯甲腈酶 A 和 B 的 K(m)值分别为 6.7 mM 和 4.5 mM。两种酶都能降解对甲苯基腈、4-氰基吡啶和对氯苯甲腈，但它们不能攻击脂肪族腈或酰胺。两种酶都被巯基试剂抑制。
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