用于B→O血型改造的不同α-半乳糖苷酶的比较

Comparison of Different α-Galactosidases Used in the B to O Blood Conversion

α 半乳糖苷酶因可水解人B型红细胞表面的α 半乳糖残基 ,使B抗原结构变成O抗原结构 ,而成为B→O血型改造的工具酶 .对可能具有酶解B抗原活性的 3种α 半乳糖苷酶 ,即来源于大豆、咖啡豆和人的α 半乳糖苷酶的结构和功能进行了比较研究 .首先 ,利用序列分析工具对 3种酶蛋白的一级结构和特性进行了比较 ;随后 ,将编码大豆和人的α 半乳糖苷酶的cDNA克隆入毕赤酵母中进行表达 ,对筛选所得表达菌株进行诱导培养 ,并从培养上清中纯化重组的大豆和人α 半乳糖苷酶 ;分别测定大豆、咖啡豆和人α 半乳糖苷酶的生物化学性质以及它们的底物特异性 ;最后 ,以纯化的重组酶对人B型红细胞进行酶解 ,并测定酶解后红细胞的结构与功能 .结果表明 ,人源的α 半乳糖苷酶不适于酶解B抗原 ,而大豆来源的α 半乳糖苷酶不仅可作为B→O血型改造的工具酶 ,而且比咖啡豆来源的α

α-Galactosidase could hydrolyze the α galactose residues on the surface of human blood type B erythrocyte, resulting in the conversion of the B antigen to O antigen. The peptide sequences of α galactosidases from soybean, coffee bean and human were aligned and analyzed with software of Lasergene. For further study, the cDNA encoding α galactosidases of soybean and human were expressed in pichia pastoris, respectively. The recombinant α galactosidase from soybean was purified by cation exchange chromatography while that from human was precipitated with ammonium sulfate and fractionated by gel filtration. Subsequently, the biochemical properties of each enzyme were assayed. The efficiency of different α galactosidase to hydrolyze the α galactose residues from human blood B erythrocyte were also compared. All the results demonstrated that the recombinant α galactosidase from soybean was more competent for B to O blood