title: Batch affinity adsorption of His-tagged proteins with EDTA-based chitosan.
abstract: Affinity adsorption purification of hexahistidine-tagged (His-tagged) proteins using EDTA-chitosan-based adsorption was designed and carried out. Chitosan was elaborated with ethylenediaminetetraacetic acid (EDTA), and the resulting polymer was characterized by FTIR, TGA, and TEM. Different metals including Ni(2+), Cu(2+), and Zn(2+) were immobilized with EDTA-chitosan, and their capability to the specific adsorption of His-tagged proteins were then investigated. The results showed that Ni(2+)-EDTA-chitosan and Zn(2+)-EDTA-chitosan had high affinity toward the His-tagged proteins, thus isolating them from protein mixture. The target fluorescent-labeled hexahistidine protein remained its fluorescent characteristic throughout the purification procedure when Zn(2+)-EDTA-chitosan was used as a sorbent, wherein the real-time monitor was performed to examine the immigration of fluorescent-labeled His-tagged protein. Comparatively, Zn(2+)-EDTA-chitosan showed more specific binding ability for the target protein, but with less binding capacity. It was further proved that this purification system could be recovered and reused at least for 5 times and could run on large scales. The presented M(2+)-EDTA-chitosan system, with the capability to specifically bind His-tagged proteins, make the purification of His-tagged proteins easy to handle, leaving out fussy preliminary treatment, and with the possibility of continuous processing and a reduction in operational cost in relation to the costs of conventional processes.
authors.affiliation: College of Food Science and Biotechnology, Zhejiang Gongshang University, Hangzhou, 310018, People's Republic of China.
authors.forename: Weiwei
authors.initials: W
authors.lastname: Hua
authors.affiliation: College of Food Science and Biotechnology, Zhejiang Gongshang University, Hangzhou, 310018, People's Republic of China.
authors.forename: Yimin
authors.initials: Y
authors.lastname: Lou
authors.affiliation: 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.affiliation: College of Food Science and Biotechnology, Zhejiang Gongshang University, Hangzhou, 310018, People's Republic of China.
authors.forename: Zhixian
authors.initials: Z
authors.lastname: Cheng
authors.affiliation: College of Food Science and Biotechnology, Zhejiang Gongshang University, Hangzhou, 310018, People's Republic of China.
authors.forename: Xingwen
authors.initials: X
authors.lastname: Gong
authors.affiliation: College of Food Science and Biotechnology, Zhejiang Gongshang University, Hangzhou, 310018, People's Republic of China. huangjy@zjgsu.edu.cn.
authors.forename: Jianying
authors.initials: J
authors.lastname: Huang
mesh_terms: D000327:Adsorption; D002614:Chelating Agents; D048271:Chitosan / Q000737:chemistry*; D002846:Chromatography, Affinity / Q000191:economics / Q000379:methods*; D004492:Edetic Acid; D004591:Electrophoresis, Polyacrylamide Gel; D006639:Histidine / Q000737:chemistry*; D008670:Metals; D009842:Oligopeptides / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry / Q000302:isolation & purification*
chemical_list: D002614:Chelating Agents; C471213:His-His-His-His-His-His; D008670:Metals; D009842:Oligopeptides; D011506:Proteins; D006639:Histidine; D048271:Chitosan; D004492:Edetic Acid
keywords: Adsorption; Chitosan; His-tagged protein; Purification; Reuse