Combination of aqueous two-phase flotation and inverse transition cycling: Strategies for separation and purification of recombinant β-glucosidase from cell lysis solution
| Autor: | Chunmei Li, Yun Wang, Yunfeng Cai, Juan Han, Lei Wang, Jiacong Wu, Sihan Fang, He Xingchen |
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| Rok vydání: | 2020 |
| Předmět: |
chemistry.chemical_classification
Chromatography Lysis Aqueous solution Recombinant Fusion Proteins beta-Glucosidase General Medicine Polymer Analytical Chemistry law.invention Enzyme Adsorption chemistry law Phase (matter) Recombinant DNA Escherichia coli Denaturation (biochemistry) Peptides Food Science |
| Zdroj: | Food chemistry. 373 |
| ISSN: | 1873-7072 |
| Popis: | This work was developed to solve the problems of the restriction of non-specific adsorption and time-dependent denaturation in the purification of recombinant proteins by multistage chromatographic procedures. A novel purification method (ATPF-ITC) which combining aqueous two-phase flotation (ATPF) with inverse transition cycling (ITC) was established and used to efficiently purify recombinant β-glucosidase (GLEGB) from cell lysis solution. First, GLEGB would preferentially adsorb on the nitrogen bubble interface relied on the hydrophobic property of the graphene-binding (GB) tag and enter into the top phase of ATPF. Second, GLEGB was achieved further purification by one-round ITC method based on the thermosensitive of the elastin-like polypeptide (ELP) tag. Consequently, the enzymatic activity recovery of GLEGB was 124.92% ± 0.83%, and the purification factor reached 24.26 ± 0.22. The purification results remained stable after six polymer cycles, and the process of ATPF-ITC had no negative effect on the structure of recombinant protein. |
| Databáze: | OpenAIRE |
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