申请人:The Mount Sinai School of Medicine of the City University of New York
公开号:US05580757A1
公开(公告)日:1996-12-03
The present invention involves the production of large quantities of human .alpha.-Gal A by cloning and expressing the .alpha.-Gal A coding sequence in eukaryotic host cell expression systems. The eukaryotic expression systems, and in particular the mammalian host cell expression system described herein provide for the appropriate cotranslational and posttranslational modifications required for proper processing, e.g., glycosylation, phosphorylation, etc. and sorting of the expression product so that an active enzyme is produced. In addition, the expression of fusion proteins which simplify purification is described. Using the methods described herein, the recombinant .alpha.-Gal A is secreted by the engineered host cells so that it is recovered from the culture medium in good yield. The .alpha.-Gal A produced in accordance with the invention may be used, but is not limited to, in the treatment in Fabry Disease; for the hydrolysis of .alpha.-galactosyl residues in glycoconjugates; and/or for the conversion of the blood group B antigen on erythrocytes to the blood group O antigen.
本发明涉及通过在真核细胞宿主细胞表达系统中克隆和表达α-半乳糖苷酶(A-Gal A)编码序列来生产大量人类A-Gal A的方法。真核表达系统,特别是在哺乳动物宿主细胞表达系统中,提供了适当的共翻译和后翻译修饰,例如糖基化,磷酸化等,以及对表达产物的分类,使得活性酶得以产生。此外,本发明还描述了简化纯化的融合蛋白的表达。使用本发明所描述的方法,重组A-Gal A由工程宿主细胞分泌,因此可以从培养基中获得良好的产量。根据本发明生产的A-Gal A可用于治疗法布里病(Fabry Disease)、水解糖基结合物中的α-半乳糖苷残基,以及将红细胞上的B型血型抗原转化为O型血型抗原等。