转化血型用蕃茄α－D－半乳糖苷酶的分离、纯化及理化性质  

PURIFICATION AND CHARACTERIZATION OF α-D-GALACTOSIDASE FROM TOMATO

作　　者：李兴德 路英华[1] 

机构地区：[1]中国学科院成都生物研究所,华东师范大学生物系

出　　处：《应用与环境生物学报》1996年第4期369-375,共7页Chinese Journal of Applied and Environmental Biology

基　　金：国家自然科学基金;中国科学院成部地奥科学基金

摘　　要：成熟蕃茄匀浆后，经硫酸铵盐析，ＤＥＡＥ－ＳｅｐｈａｄｅｘＡ－５０离子交换层析，ＳｅｐａｄｅｘＧ－１００凝胶过滤和Ｍｅｌｉｂｉｏｓｅ－Ａｇａｒｏｓｅ亲和层析，获得了α－Ｄ－半乳糖苷酶（Ｃ．Ｅ．３．２．１．１１）。酶制剂经ＰＡＧＥ检测为一条带；ＳＤＳ－Ｇ－ＰＡＧＥ测得酶Ｍｒ为３４０００；比活力５２．９Ｕ／ｍｇ·；提纯倍数为５２９０１产率为４５％．酶专－催化以α－Ｄ－半乳糖为末端ａ－（１，３）连接的糖苷键，以ＰＮＰＧ（对硝基苯－α－Ｄ－半乳糖昔）为废物，酶催化反应的Ｋｍ＝０．１１ｍｍｏｌ／Ｌ，Ｖｍａｘ为６７μｍｏｌ·ｍｇ１－·ｍｉｎ－１．ｔ稳定范是０～３５℃；ＰＨ稳定范围是４．０～７．０．最适ｐＨ为５．１．半乳糖是酶的竞争性抑制剂；Ｃｕ２＋、Ｚｎ２＋、Ｍｎ２＋、Ｆｅ３＋、Ａｇ＋和ＥＤＴＡ对酶活性无影响．纯酶制剂可作为Ｂ型血向Ｏ型血转化的工具酶液．Tomato(Lycoporiscoum ) was selected as raw material for purilyins α-D-galactosidase.Purified tomatoα-D-galactosidase was obtained by (NH4)2SO4 treatment,DEAE-Sephadex A-50 ion-exchange chromatography,Sephadex G- 100 gel filtration and Melibiose-agnrose affinity chromatography. The results showed thata 5 290 fold purification of enzyme was obtained, its recovery rate was 45%,and the specific activity was52. 9 U/mg. pro.The enzyme's M, was 34 000 by SDS-G-PAGE. The tomato α-D-galactosidase was stableat pH 4. 0￣ 7. 0 and tempetature 0 ￣ 35℃. The enzyme hydrolysed substrate PNPG with the Km 0. 11mmol/L,V,max = 67 μmol· mg-1· min-1,optimum pH 5. 1. The α-D-galactose is a competitive inhibitor ofthe enzyme. Metal ions CU2+ .Zn2+ .Mn2+ .Fe3+ .Ag+ and EDTA have no effects upon enzymology activity.

关 键 词：番茄 血型转化 α-D半乳糖苷酶 分离 理化性质 

分 类 号：S641.201[农业科学—蔬菜学] R457.11[农业科学—园艺学]