title: Large-scale production of soluble recombinant amyloid-β peptide 1-42 using cold-inducible expression system.
abstract: Amyloid-β peptide 1-42 (Aβ(1-42)), the predominant form in senile plaques, plays important roles in the pathogenesis of Alzheimer's disease. Because Aβ(1-42) has aggregation-prone nature, it has been difficult to produce in a soluble state in bacterial expression systems. In this study, we modified our expression system to increase the soluble fraction of Aβ(1-42) in Escherichia coli (E. coli) cells. The expression level and solubility of recombinant Aβ(1-42) induced at the low temperature (16°C) is highly increased compared to that induced at 37°C. To optimize expression temperature, the coding region of Aβ(1-42) was constructed in a pCold vector, pCold-TF, which has a hexahistidine-tagged trigger factor (TF). Recombinant Aβ(1-42) was expressed primarily as a soluble protein using pCold vector system and purified with a nickel-chelating resin. When the toxic effect of recombinant Aβ(1-42) examined on human neuroblastoma SH-SY5Y cells, the purified Aβ(1-42) induced cell toxicity on SH-SY5Y cells. In conclusion, the system developed in this study will provide a useful method for the production of aggregation prone-peptide such as Aβ(1-42).
authors.affiliation: Lee Gil Ya Cancer and Diabetes Institute, Gachon University, Incheon 406-799, Republic of Korea.
authors.forename: Eun-Kyung
authors.initials: EK
authors.lastname: Kim
authors.forename: Jeong Chan
authors.initials: JC
authors.lastname: Moon
authors.forename: Jeong Mi
authors.initials: JM
authors.lastname: Lee
authors.forename: Min Seop
authors.initials: MS
authors.lastname: Jeong
authors.forename: Choongseob
authors.initials: C
authors.lastname: Oh
authors.forename: Sung-Min
authors.initials: SM
authors.lastname: Ahn
authors.forename: Yung Joon
authors.initials: YJ
authors.lastname: Yoo
authors.forename: Ho Hee
authors.initials: HH
authors.lastname: Jang
mesh_terms: D000595:Amino Acid Sequence; D016229:Amyloid beta-Peptides / Q000096:biosynthesis* / Q000235:genetics; D045744:Cell Line, Tumor; D002470:Cell Survival; D002846:Chromatography, Affinity / Q000379:methods; D003080:Cold Temperature; D004926:Escherichia coli / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics*; D006801:Humans; D009447:Neuroblastoma / Q000473:pathology; D016366:Open Reading Frames; D010446:Peptide Fragments / Q000096:biosynthesis* / Q000235:genetics; D059748:Proteolysis; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics; D012995:Solubility; D014170:Transformation, Genetic
chemical_list: D016229:Amyloid beta-Peptides; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; C075222:amyloid beta-protein (1-42)