title: A new method for purification of recombinant human alpha-synuclein in Escherichia coli.
abstract: alpha-Synuclein (AS), a major component of Lewy body in Parkinson's disease patients, exists as a natively unfolded protein in physiological buffer. We recently found that the overexpressed AS in Escherichia coli bearing the cloned AS cDNA with no signal sequence was actually located inside the periplasm, but not in the cytoplasm as generally recognized. Therefore, a new protocol for preparing recombinant AS has been developed with only two steps: (1) osmotic shock for release of AS-containing periplasm fraction and (2) ion-exchange chromatography for further purification of AS. By using plasmids and E. coli strains commonly used the new protocol is much more convenient, faster, and cheaper compared to the current methods established since 1994. About 80 mg AS with 95% purity can be regularly prepared from a 1L culture in 3 days.
authors.affiliation: The National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing.
authors.forename: Chunjuan
authors.initials: C
authors.lastname: Huang
authors.forename: Guoping
authors.initials: G
authors.lastname: Ren
authors.forename: Hui
authors.initials: H
authors.lastname: Zhou
authors.forename: Chih-chen
authors.initials: CC
authors.lastname: Wang
mesh_terms: D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D002942:Circular Dichroism; D004926:Escherichia coli / Q000235:genetics*; D006801:Humans; D008854:Microscopy, Electron; D008970:Molecular Weight; D046912:Multiprotein Complexes / Q000096:biosynthesis / Q000648:ultrastructure; D009997:Osmotic Pressure; D019897:Periplasm / Q000737:chemistry; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification*; D013050:Spectrometry, Fluorescence
chemical_list: D046912:Multiprotein Complexes; D011994:Recombinant Proteins