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Design of Affinity Tags for One‐Step Protein Purification from Immobilized Zinc Columns
Author(s) -
Pasquinelli Richard S.,
Shepherd Rex E.,
Koepsel Richard R.,
Zhao Adam,
Ataai Mohammad M.
Publication year - 2000
Publication title -
biotechnology progress
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 129
eISSN - 1520-6033
pISSN - 8756-7938
DOI - 10.1021/bp990139h
Subject(s) - zinc , chromatography , chemistry , affinity chromatography , protein purification , biochemistry , organic chemistry , enzyme
Affinity tags are often used to accomplish recombinant protein purification using immobilized metal affinity chromatography. Success of the tag depends on the chelated metal used and the elution profile of the host cell proteins. Zn(II)‐iminodiacetic acid (Zn(II)‐IDA) may prove to be superior to either immobilized copper or nickel as a result of its relatively low binding affinity for cellular proteins. For example, almost all Escherichia coli proteins elute from Zn(II)‐IDA columns between pH 7.5 and 7.0 with very little cellular protein emerging at pH values lower than 7.0. Thus, a large portion of the Zn(II)‐IDA elution profile may be free of contaminant proteins, which can be exploited for one‐step purification of a target protein from raw cell extract. In this paper we have identified several fusion tags that can direct the elution of the target protein to the low background region of the Zn(II)‐IDA elution profile. These tags allow targeting of proteins to different regions of the elution profile, facilitating purification under mild conditions.