title: A multipurpose fusion tag derived from an unstructured and hyperacidic region of the amyloid precursor protein.
abstract: Expression and purification of aggregation-prone and disulfide-containing proteins in Escherichia coli remains as a major hurdle for structural and functional analyses of high-value target proteins. Here, we present a novel gene-fusion strategy that greatly simplifies purification and refolding procedure at very low cost using a unique hyperacidic module derived from the human amyloid precursor protein. Fusion with this polypeptide (dubbed FATT for Flag-Acidic-Target Tag) results in near-complete soluble expression of variety of extracellular proteins, which can be directly refolded in the crude bacterial lysate and purified in one-step by anion exchange chromatography. Application of this system enabled preparation of functionally active extracellular enzymes and antibody fragments without the need for condition optimization.
authors.affiliation: Laboratory of Protein Synthesis and Expression, Institute for Protein Research, Osaka University, Suita, Osaka, Japan.
authors.forename: Takeshi
authors.initials: T
authors.lastname: Sangawa
authors.forename: Sanae
authors.initials: S
authors.lastname: Tabata
authors.forename: Kei
authors.initials: K
authors.lastname: Suzuki
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Saheki
authors.forename: Keiji
authors.initials: K
authors.lastname: Tanaka
authors.forename: Junichi
authors.initials: J
authors.lastname: Takagi
mesh_terms: D000595:Amino Acid Sequence; D016564:Amyloid beta-Protein Precursor / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification; D003001:Cloning, Molecular / Q000379:methods; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D058849:Protein Refolding; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D012995:Solubility
chemical_list: D016564:Amyloid beta-Protein Precursor; D011993:Recombinant Fusion Proteins