摘要
三角酵母 (Trigonopsisvariabilis)的D 氨基酸氧化酶 (D aminoacidoxidase ,DAO ,EC1 .4.3.3)是一种胞内酶 ,完整细胞并不呈现酶促活性。为了获得三角酵母细胞的较高表观活力 ,需对细胞进行处理以改变壁和膜对反应底物和产物的通透性。研究中比较了冻融、丙酮、丁醇和十六烷基三甲基溴化铵 (Cetyltrimethylammoniumbro mide ,CTAB)等理化因子的透性化效率 ,并对丙酮的透性化条件进行了优化。证明丙酮的透性化作用与溶剂浓度、保温时间和温度有关。 30 %～ 35 %丙酮浓度 ,4～ 2 8℃保温 ,可得到最大酶活力。透性化所需时间极短 ,在 5min内即可完成。同时 ,透性化细胞中的DAO比无细胞抽提液中的酶对热更为稳定。用丙酮透性化细胞作为生物催化剂可有效地转化头孢菌素C(CephalosporinC ,CPC)为戊二酰 7ACA(Glutaryl 7 ACA ,GL 7ACA)。
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℃ and 28℃. 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.
出处
《生物工程学报》
CAS
CSCD
北大核心
2001年第1期73-77,共5页
Chinese Journal of Biotechnology
基金
国家"九五"科技攻关项目!(96 1 0 2 0 2 0 1 )
上海市科学技术发展基金!(98431 91 35)资助&&
