摘要
D-海因酶是海因酶法制备D-氨基酸的关键酶。利用Burkholderic cepecia1003菌发酵产酶,所得海因酶纯化后,以Eupergit C250L为载体进行共价固定化。分别考察了酶液蛋白浓度、固定化时间对蛋白固定量和酶活回收率的影响以及固定化前后海因酶催化性质的变化。结果表明:较高的酶液蛋白浓度和较长的固定化时间均有助于改善海因酶的固定化效果;固定化可显著提高海因酶的最适作用温度,但对其最适作用pH影响不大;固定化后海因酶对D,L-BH和MH的米氏常数均有较大幅度的降低。固定化酶反应器的实验表明:40℃下,底物(D,L-BH)1.0 g.L-1,体积流速1.0 mL.min-1,经21 h转化,产物N-Phe质量浓度可达0.47 g.L-1,转化率达43.21%。
D-hydantoinase was a key enzyme, which was used to prepare D-phenylalanine. The D- hydantoinase from Burkholderic cepecia 1003 was purified and immobilized to Eupergit C250L by covalent bonds. The effects of protein concentration of enzyme solution, reaction time on yield of immobilizing protein and enzyme activity were measured. The variety of catalyze capability of free and immobilized D-hydantoinase were investigated. The results showed that higher protein concentration of enzyme solution and longer reaction time could improve the immobilization of hydantoinase. Compared to the free enzyme, the immobilized enzyme exhibited higher reaction temperature but less effection to pH. The measured Km value of immobilized enzyme was lower than that of free enzyme when acting on both D, L-BH and MH. Packed-bod reactor of immobilized enzyme had an optimal conditions of 40 ℃, a flow rate of 1.0 mL·min^- 1, concentration of substrate （ D, L- BH） was 1.0 g·L^-1. The concentration of N-Phe reached 0.47 g·L^-1 and the rate of conversion was 43.21% after 21 h.
出处
《生物加工过程》
CAS
CSCD
2006年第4期41-45,共5页
Chinese Journal of Bioprocess Engineering
基金
国家973项目(No.2003CB7160004)
