Nitrilase of Rhodococcus rhodochrous J1 -: Conversion into the active form by subunit association
Nitrilase of Rhodococcus rhodochrous J1 -: Conversion into the active form by subunit association
复制标题
DOI：
10.1046/j.1432-1327.2000.00983.x
复制
发表时间：
2000-01-01
期刊：
影响因子：
--
通讯作者：
Gekko, K
中科院分区：
文献类型：
--
作者：
Nagasawa, T;Wieser, M;Gekko, K
含腈酶的红球菌J1静息细胞将丙烯腈和苯腈转化为相应的酸，但纯化后的腈酶只水解苯腈，不水解丙烯腈。纯化后的酶对丙烯腈的活性可通过10 mm苯腈预孵育恢复，但不能通过与丙烯腈等脂肪族腈预孵育恢复。光散射实验表明，与苯腈预孵育导致无活性、纯化的同二聚体80 kda酶组装成活性的410 kda聚集体，该聚集体被认为是一个十聚体。此外，在对各种盐和有机溶剂进行透析后，酶与活化的关联达到了，在10%饱和硫酸铵和50% （v/v）甘油以及在更高的温度或酶浓度下进行预孵育时，回收率最高。
Nitrilase-containing resting cells of Rhodococcus rhodochrous J1 converted acrylonitrile and benzonitrile to the corresponding acids, but the purified nitrilase hydrolyzed only benzonitrile, and not acrylonitrile. The activity of the purified enzyme towards acrylonitrile was recovered by preincubation with 10 mm benzonitrile, but not by preincubation with aliphatic nitriles such as acrylonitrile. It was shown by light-scattering experiments, that preincubation with benzonitrile led to the assembly of the inactive, purified and homodimeric 80-kDa enzyme to its active 410-kDa aggregate, which was proposed to be a decamer. Furthermore, the association concomitant with the activation was reached after dialysis of the enzyme against various salts and organic solvents, with the highest recovery reached at 10% saturated ammonium sulfate and 50% (v/v) glycerol, and by preincubation at increased temperatures or enzyme concentrations.