title: Cloning, soluble expression, rapid purification and characterization of human Cofilin1.
abstract: Cofilin1 is an actin-binding protein that plays a critical role in the regulation of actin cytoskeleton and consequently affects various physiological processes. In this study, the human Cofilin1 cDNA was cloned into the expression vector pET-28a(+) with a 6 × His tag and expressed as soluble protein in Escherichia coli BL21(DE3). Approximately 78 mg of Cofilin1, which showed high activity as determined by native PAGE, could be purified from each liter of LB medium by His-tag affinity chromatography and gel filtration. Further, high-titer IgG against Cofilin1 was positively detected after immunization in rabbits and the polyclonal antibodies were purified and identified. Together, this report provides the first protocol to efficiently obtain human Cofilin1 with high biological activity and immunogenicity using E. coli BL21 (DE3) expression system.
authors.affiliation: Biomedicine Research and Development Center, Guangdong Provincial Key Laboratory of Bioengineering Medicine, National Engineering Research Center of Genetic Medicine, Jinan University, Guangzhou 510632, China.
authors.forename: Jia-Xin
authors.initials: JX
authors.lastname: Lu
authors.forename: Yang-Fei
authors.initials: YF
authors.lastname: Xiang
authors.forename: Jia-Xuan
authors.initials: JX
authors.lastname: Zhang
authors.forename: Huai-Qiang
authors.initials: HQ
authors.lastname: Ju
authors.forename: Zhen-Ping
authors.initials: ZP
authors.lastname: Chen
authors.forename: Qiao-Li
authors.initials: QL
authors.lastname: Wang
authors.forename: Wei
authors.initials: W
authors.lastname: Chen
authors.forename: Xin-Lei
authors.initials: XL
authors.lastname: Peng
authors.forename: Bo
authors.initials: B
authors.lastname: Han
authors.forename: Yi-Fei
authors.initials: YF
authors.lastname: Wang
mesh_terms: D000818:Animals; D000906:Antibodies / Q000276:immunology; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D051338:Cofilin 1 / Q000737:chemistry / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification*; D018076:DNA, Complementary / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D006639:Histidine / Q000235:genetics; D006801:Humans; D007114:Immunization; D009842:Oligopeptides / Q000235:genetics; D011817:Rabbits; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D012995:Solubility
chemical_list: D000906:Antibodies; D051338:Cofilin 1; D018076:DNA, Complementary; C471213:His-His-His-His-His-His; D009842:Oligopeptides; D011994:Recombinant Proteins; D006639:Histidine