title: An AIL/IL-based liquid/liquid extraction system for the purification of His-tagged proteins.
abstract: A sorbent based on affinity ionic liquid (AIL), triazacyclononane-ionic liquid, was synthesized, characterized, and applied to the extraction of histidine (His)-tagged proteins from aqueous buffer to ionic liquid (IL) phase. The adsorbed His-tagged proteins could be back-extracted from the IL phase to the aqueous buffer with an imidazole solution. The specific binding of His-tagged proteins with AIL/IL could be affected by a few factors including the ionic strength and coordinated metal ions. In the case of His-tagged enhanced green fluorescent protein (EGFP), the maximum binding capacity of Cu(2+)-AIL/IL reached 2.58 μg/μmol under the optimized adsorption conditions. The eluted His-tagged EGFP kept fluorescent and remained active through the purification process. Moreover, a tandem extraction process successively using Cu(2+)-AIL/IL and Zn(2+)-AIL/IL systems was developed, which was proven very efficient to obtain the ultimate protein with a purity of about 90 %. An effective reclamation method for the AIL/IL extraction system was further established. The sorbent could be easily regenerated by removing metal ions with EDTA and the followed reimmobilization of metal ions. Easy handling of the presented M(2+)-AIL/IL system and highly specific ability to absorb His-tagged proteins make it attractive and potentially applicable in biomolecular separation.
authors.affiliation: Key Laboratory of Fruits and Vegetables Postharvest and Processing Technology Research of Zhejiang Province, College of Food Science and Biotechnology, Zhejiang Gongshang University, Hangzhou, 310018, People's Republic of China.
authors.forename: Weiyuan
authors.initials: W
authors.lastname: Xu
authors.forename: Huazhen
authors.initials: H
authors.lastname: Cao
authors.forename: Guangwei
authors.initials: G
authors.lastname: Ren
authors.forename: Hujun
authors.initials: H
authors.lastname: Xie
authors.forename: Jianying
authors.initials: J
authors.lastname: Huang
authors.forename: Shijun
authors.initials: S
authors.lastname: Li
mesh_terms: D000327:Adsorption; D001372:Aza Compounds / Q000737:chemistry; D052578:Ionic Liquids / Q000737:chemistry; D059625:Liquid-Liquid Extraction / Q000379:methods*; D010880:Piperidines / Q000737:chemistry; D011506:Proteins / Q000737:chemistry / Q000302:isolation & purification*
chemical_list: D001372:Aza Compounds; D052578:Ionic Liquids; D010880:Piperidines; D011506:Proteins; C482777:triazacyclononane