Purification of microbial b-galactosidase from Kluyveromyces fragilis by bioaffinity partitioning

Revista de Microbiologia

Universidade Estadual de Campinas, unicamp, Campinas, Brasil¹  Universidade Estadual de Campinas, Campinas, Brasil¹  Silva, Maria Estela da¹  Franco, Telma Teixeira¹ 

关键词: : b-galactosidase;    aqueous two-phase systems;    protein purification;    downstream-processing;    affinity;   

DOI  :  10.1590/S0001-37141999000400006

学科分类：农业科学（综合）

来源: Sociedade Brasileira de Microbiologia / Brazilian Society for Microbiology

【 摘 要 】

This work investigated the partitioning of b-galactosidase from Kluyveromyces fragilis in aqueous two-phase systems (ATPS) by bioaffinity. PEG 4000 was chemically activated with thresyl chloride, and the biospecific ligand p-aminophenyl 1-thio-b-D-galactopyranoside (APGP) was attached to the activated PEG 4000. A new two-step method for extraction and purification of the enzyme b-galactosidase from Kluyveromyces fragilis was developed. In the first step, a system composed of 6% PEG 4000-APGP and 8% dextran 505 was used, where b-galactosidase was strongly partitioned to the top phase (K = 2,330). In the second step, a system formed of 13% PEG-APGP and 9% phosphate salt was used to revert the value of the partition coefficient of b-galactosidase (K = 2 x 10-5) in order to provide the purification and recovery of 39% of the enzyme in the bottom salt-rich phase.

【 授权许可】

Unknown