Immobilization of $\beta$-Galactosidase from E. coli K-12 CHS36 Using Tannin - Activated Cellulose Beads

$\beta$-Galactosidase의 Tannin 활성화 섬유소 입자에의 고정화

$\beta$-Galactosidase($\beta$-D-galactoside galactohydrolase, E.C. 3. 2. 1. 23) from E. coli K-12 CSH 36 was immobilized on porous cellulose beads which were previously activated with tannin and p-benzoquinone. Their general properties and applicational possibities were investigated. The most effective, enzyme immobilization was obtained when tannin and p-benzoquinone, pH 11.0, were used together as activation reagents and a period of 6 hours of activation. The optimum pH of $\beta$-galactosidase was 5.5 for free enzyme and 6. 0 for the immobilized enzyme, the optimum temperatures for native and immobilized enzyme were both $50^{\circ}C$. Kms of native $\beta$-galactosidase and immobilized enzyme for ONPG(o-nitrophenyl galactopyranoside) were about $4.0{\times}10^(-4)M$ and $7.5{\times}10^(-4)M$, respectively. In the case of tannin : p-benzoquinone activated cellulose beads, the immobilized enzyme retained over 80% of the initial enzyme activity after 20 runs, which is very promising result far a possible industrial application.

Tannin 및 p-benzoquinone으로 활성화시킨 다공성 섬유소 입자를 사용하여 대장균 변이주(E. coli K-12 CSH36)로부터 추출한 $\beta$-galactosidase를 고정화시켜 효소학적 성질과 그 응용성에 대하여 연구하였다. $\beta$-galactosidase는 pH 11.0으로 맞춘 tannin과 p-benzoquinone을 사용하여 6시간 동안 활성화시킨 섬유소 입자담체에 고정화 시켰을 때 그 잔여 활성도가 가장 높았다. $\beta$-galactosidase의 최적작용 pH는 5.5였으나 고정화 시켰을 때 pH 6.0으로 변화하였으며 최적작용 온도는 고정화 전이나 후에도 일정하였다. $\beta$-galactosidase의 Km 값은 $4.0{\times}10^(-4)M$으로 나타났으며 고정화 효소의 경우에는 $7.5{\times}10^(-4)M$이었다. 고정화 효소의 재사용성을 검토한 결과, 담체 활성화 물질로 tannin과 p-benzoquinone 용액을 사용하였을 때 20히 사용 후에 초기 효소 활성도의 80% 이상이 유지되는 결과를 나타내었다.