[用于提高 D-氨基酸氧化酶表观活性的可变三角酵母 FA10 细胞的透化处理]

[Permeabilization of yeast Trigonopsis variabilis FA10 cells for enhancing apparent activity of D-amino acid oxidase].

作者信息

Zhu T B, Chen J, Zhang Y F, Yang Y L, Jiao R S

机构信息

Shanghai Institute of Plant Physiology, Chinese Academy of Sciences, Shanghai 200032, China.

出版信息

Sheng Wu Gong Cheng Xue Bao. 2001 Jan;17(1):73-7.

PMID:11330193

Abstract

D-amino acid oxidase(DAO) is an intracellular enzyme in Trigonopsis variabilis FA10. The whole cells of yeast exhibited very low DAO activity. Various treatment with physical and chemical agents, such as, freezing and thawing, acetone, butanol and cetyltrimethylammoinium bromide altered membrane permeability and increased cellular DAO activity. It was demonstrated that the performance of acetone was dependent on the concentration of the solvent, the incubation time and the temperature. Maximum enzyme activity of the cells was achieved with 30-35% acetone, between 4 degrees C and 28 degrees C. The process was very quick and permeabilization occurred in 5 minutes or less. On the other hand, the thermostability of DAO in permeabilized cells was higher than in cell extract. It was proved that Cephalosporin C could be effectively converted to glutaryl-7ACA by permeabilized Trigonopsis variabilis cells with 83% yield.

摘要

D-氨基酸氧化酶（DAO）是可变三角酵母FA10中的一种细胞内酶。酵母全细胞表现出非常低的DAO活性。用物理和化学试剂进行的各种处理，如冻融、丙酮、丁醇和十六烷基三甲基溴化铵，改变了膜通透性并提高了细胞DAO活性。结果表明，丙酮的作用取决于溶剂浓度、孵育时间和温度。在4℃至28℃之间，用30-35%的丙酮可使细胞达到最大酶活性。该过程非常迅速，在5分钟或更短时间内即可实现通透化。另一方面，通透化细胞中DAO的热稳定性高于细胞提取物中的热稳定性。事实证明，用可变三角酵母通透化细胞可将头孢菌素C有效转化为戊二酰-7-氨基头孢烷酸，产率为83%。