title: An epitope tag derived from human transcription factor IIB that reacts with a polyol-responsive monoclonal antibody.
abstract: Polyol-responsive monoclonal antibodies (PR-mAbs) provide a strategy to purify active, nondenatured proteins by a single-step immunoaffinity chromatography procedure. The high affinity interaction between these antibodies and the antigen can be dissociated in the presence of a nonchaotropic salt and a low molecular weight polyhydroxylated compound (polyol). The epitope for PR-mAb IIB8 is located near the N-terminus of the human transcription factor IIB (TFIIB). The epitope is an eight amino acid sequence, TKDPSRVG, that can be fused to a desired protein for use as a purification tag. This epitope tag (termed hIIB) was fused to the C-terminus of green fluorescent protein (GFP). An additional GFP fusion protein utilized another version of hIIB containing a point mutation at position two. These fusion proteins, expressed in Escherichia coli, allowed successful separation of the desired protein in a single chromatographic step. This strategy extends PR-mAb gentle-release purification to numerous expressed proteins.
authors.affiliation: McArdle Laboratory for Cancer Research, University of Wisconsin-Madison, Madison, WI 53706, USA. sjduellman@wisc.edu
authors.forename: Sarah J
authors.initials: SJ
authors.lastname: Duellman
authors.forename: Nancy E
authors.initials: NE
authors.lastname: Thompson
authors.forename: Richard R
authors.initials: RR
authors.lastname: Burgess
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000378:metabolism; D002846:Chromatography, Affinity / Q000379:methods*; D018604:Epitope Mapping; D000939:Epitopes; D049452:Green Fluorescent Proteins; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D011108:Polymers / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D035581:Transcription Factor TFIIB / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D000939:Epitopes; D008164:Luminescent Proteins; D011108:Polymers; D011993:Recombinant Fusion Proteins; D035581:Transcription Factor TFIIB; C024617:polyol; D049452:Green Fluorescent Proteins