Recombinant Protein Purification using Composite Polyacrylamide-Nanocrystalline Cryogel Monolith Column and Carbohydrate- Binding Module Family ۶۴ as Affinity Tag
- سال انتشار: 1401
- محل انتشار: مجله گزارش های بیوشیمی و زیست شناسی مولکولی، دوره: 11، شماره: 2
- کد COI اختصاصی: JR_RBMB-11-2_008
- زبان مقاله: انگلیسی
- تعداد مشاهده: 178
نویسندگان
Department of Medical Biotechnology, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran, Iran.
Department of Medical Biotechnology, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran, Iran.
Department of Biotechnology, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran.
Department of Medical Biotechnology, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran, Iran.
چکیده
Background: In the field of recombinant protein production, downstream processing, especially protein purification, is critical and often the most expensive step. Carbohydrate binding module ۶۴ (CBM۶۴) was shown in ۲۰۱۱ to bind efficiently to a broad range of cellulose materials. Methods: In this study, we developed a protein purification method using nanocrystalline cellulose embedded in a polyacrylamide monolith cryogel and CBM۶۴ affinity tag linked by intein to PD۱ as a model protein. The CBM۶۴-Intein-PD۱ gene cassette was expressed in E. coli. Following cell lysis, CBM۶۴-Intein-PD۱ protein bound to the monolith PA-NCC cryogel. After washing and reducing the pH from ۸.۰ to ۶.۵, the intein underwent self-cleavage, resulting in the release and elution of pure PD۱ protein. Results: The synthesized monolith column had a porous structure with an average pore size of ۳۰ μm and a maximum binding capacity of ۴۹۷ μg per gram of dried column. The yield of this purification method was ۸۴%, while the yield of the His tag-acquired CBM۶۴-Intein-PD۱ method was ۸۹%. Conclusions: We used cellulose as support for affinity chromatography, which can be used as a cost-effective method for protein purification.کلیدواژه ها
Affinity tag, CBM۶۴, PD۱, Protein purification.اطلاعات بیشتر در مورد COI
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