title: Design of affinity tags for one-step protein purification from immobilized zinc columns.
abstract: 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.
authors.affiliation: Department of Chemical Engineering, Center for Biotechnology and Bioengineering, University of Pittsburgh, Pittsburgh, Pennsylvania 15219, USA.
authors.forename: R S
authors.initials: RS
authors.lastname: Pasquinelli
authors.forename: R E
authors.initials: RE
authors.lastname: Shepherd
authors.forename: R R
authors.initials: RR
authors.lastname: Koepsel
authors.forename: A
authors.initials: A
authors.lastname: Zhao
authors.forename: M M
authors.initials: MM
authors.lastname: Ataai
mesh_terms: D000345:Affinity Labels; D000595:Amino Acid Sequence; D001483:Base Sequence; D001709:Biotechnology; D002846:Chromatography, Affinity / Q000379:methods*; D015195:Drug Design; D004926:Escherichia coli / Q000235:genetics; D009838:Oligodeoxyribonucleotides / Q000235:genetics; D015202:Protein Engineering; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification*; D015032:Zinc
chemical_list: D000345:Affinity Labels; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D015032:Zinc