title: Purification and on-column refolding of a single-chain antibody fragment against rabies virus glycoprotein expressed in Escherichia coli.
abstract: An anti-rabies virus single-chain antibody fragment of an anti-glycoprotein with the VL-linker-VH orientation, designated scFv57RN, was successfully and conveniently prepared in this study. The scFv57RN protein was mainly expressed in inclusion bodies in Escherichia coli. After washing and purification, the inclusion bodies were finally obtained with an on-column refolding procedure. Further purification by gel exclusion chromatography was performed to remove inactive multimers. About 360 mg of final product was recovered from 1 L of bacterial culture. The final product showed a high neutralizing titer of 950 IU/mg to the CVS-11 strain as measured using the rapid fluorescent focus inhibition test. Our study demonstrated a highly efficient method to mass produce scFV57RN with activity from inclusion bodies, which may be applied in the purification of other insoluble proteins.
authors.affiliation: National Engineering Laboratory for AIDS Vaccine, School of Life Science, Jilin University, Changchun 130012, China.
authors.forename: Hualong
authors.initials: H
authors.lastname: Xi
authors.affiliation: BCHT Biotechnology Company, Changchun 130012, China.
authors.forename: Ruosen
authors.initials: R
authors.lastname: Yuan
authors.affiliation: BCHT Biotechnology Company, Changchun 130012, China.
authors.forename: Xiaoxu
authors.initials: X
authors.lastname: Chen
authors.affiliation: National Engineering Laboratory for AIDS Vaccine, School of Life Science, Jilin University, Changchun 130012, China; Key Laboratory for Molecular Enzymology and Engineering of the Ministry of Education, School of Life Science, Jilin University, Changchun 130012, China.
authors.forename: Tiejun
authors.initials: T
authors.lastname: Gu
authors.affiliation: National Engineering Laboratory for AIDS Vaccine, School of Life Science, Jilin University, Changchun 130012, China.
authors.forename: Yue
authors.initials: Y
authors.lastname: Cheng
authors.affiliation: National Engineering Laboratory for AIDS Vaccine, School of Life Science, Jilin University, Changchun 130012, China.
authors.forename: Zhuang
authors.initials: Z
authors.lastname: Li
authors.affiliation: National Engineering Laboratory for AIDS Vaccine, School of Life Science, Jilin University, Changchun 130012, China; Key Laboratory for Molecular Enzymology and Engineering of the Ministry of Education, School of Life Science, Jilin University, Changchun 130012, China.
authors.forename: Chunlai
authors.initials: C
authors.lastname: Jiang
authors.affiliation: National Engineering Laboratory for AIDS Vaccine, School of Life Science, Jilin University, Changchun 130012, China; Key Laboratory for Molecular Enzymology and Engineering of the Ministry of Education, School of Life Science, Jilin University, Changchun 130012, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Kong
authors.affiliation: National Engineering Laboratory for AIDS Vaccine, School of Life Science, Jilin University, Changchun 130012, China; Key Laboratory for Molecular Enzymology and Engineering of the Ministry of Education, School of Life Science, Jilin University, Changchun 130012, China. Electronic address: ygwu@jlu.edu.cn.
authors.forename: Yongge
authors.initials: Y
authors.lastname: Wu
mesh_terms: D004926:Escherichia coli; D015870:Gene Expression; D006023:Glycoproteins / Q000032:analysis* / Q000737:chemistry; D002479:Inclusion Bodies / Q000737:chemistry / Q000378:metabolism; D058849:Protein Refolding; D011820:Rabies virus / Q000737:chemistry*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D057127:Single-Chain Antibodies / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D014764:Viral Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry
chemical_list: D006023:Glycoproteins; D011994:Recombinant Proteins; D057127:Single-Chain Antibodies; D014764:Viral Proteins
keywords: Fermentation; On-column refolding; Purification; Rabies virus; scFv