Tags

Type your tag names separated by a space and hit enter

Gene cloning, bacterial expression, in vitro refolding, and characterization of a single-chain Fv antibody against PreS1(21-47) fragment of HBsAg.
Protein Expr Purif. 2005 Jun; 41(2):341-8.PE

Abstract

The murine monoclonal antibody 125E11 is an IgG which recognizes PreS1(21-47) fragment of large hepatitis B surface antigen. It has been successfully used for clinical detection of HBV virion in serum of hepatitis B patients. In present study, the genes of variable region in heavy chain (VH) and light chain (VL) of 125E11 have been cloned. Sequence analysis of cloned VH gene and VL gene showed that they had general characterization of immunoglobin variable region genes. According to Kabat classification, VH gene and VL gene belong to VH10 family, subgroup IIID and Vkappa family subgroup I, respectively. An expression vector of 125E11 single-chain Fv antibody fusion protein, in which VH and VL peptide were connected by a flexible linker (Gly(4)Ser)(3), was constructed. The scFv fusion protein was highly expressed in Escherichia coli mainly in inclusion body form. Using urea and pH gradient gel filtration method, the refolding of scFv was efficiently achieved. The refolding efficiency reached about 11% and 2.7 mg refolded scFv was obtained from 1L of culture. The binding activity and specificity of 125E11 scFv against PreS1(21-47)-containing antigen were also analyzed.

Authors+Show Affiliations

Key Laboratory of Proteomics, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article

Language

eng

PubMed ID

15866720

Citation

Yang, Xinxiu, et al. "Gene Cloning, Bacterial Expression, in Vitro Refolding, and Characterization of a Single-chain Fv Antibody Against PreS1(21-47) Fragment of HBsAg." Protein Expression and Purification, vol. 41, no. 2, 2005, pp. 341-8.
Yang X, Hu W, Li F, et al. Gene cloning, bacterial expression, in vitro refolding, and characterization of a single-chain Fv antibody against PreS1(21-47) fragment of HBsAg. Protein Expr Purif. 2005;41(2):341-8.
Yang, X., Hu, W., Li, F., Xia, H., & Zhang, Z. (2005). Gene cloning, bacterial expression, in vitro refolding, and characterization of a single-chain Fv antibody against PreS1(21-47) fragment of HBsAg. Protein Expression and Purification, 41(2), 341-8.
Yang X, et al. Gene Cloning, Bacterial Expression, in Vitro Refolding, and Characterization of a Single-chain Fv Antibody Against PreS1(21-47) Fragment of HBsAg. Protein Expr Purif. 2005;41(2):341-8. PubMed PMID: 15866720.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Gene cloning, bacterial expression, in vitro refolding, and characterization of a single-chain Fv antibody against PreS1(21-47) fragment of HBsAg. AU - Yang,Xinxiu, AU - Hu,Weiguo, AU - Li,Feng, AU - Xia,Hengchuan, AU - Zhang,Zuchuan, PY - 2004/12/08/received PY - 2005/02/05/revised PY - 2005/5/4/pubmed PY - 2005/11/4/medline PY - 2005/5/4/entrez SP - 341 EP - 8 JF - Protein expression and purification JO - Protein Expr. Purif. VL - 41 IS - 2 N2 - The murine monoclonal antibody 125E11 is an IgG which recognizes PreS1(21-47) fragment of large hepatitis B surface antigen. It has been successfully used for clinical detection of HBV virion in serum of hepatitis B patients. In present study, the genes of variable region in heavy chain (VH) and light chain (VL) of 125E11 have been cloned. Sequence analysis of cloned VH gene and VL gene showed that they had general characterization of immunoglobin variable region genes. According to Kabat classification, VH gene and VL gene belong to VH10 family, subgroup IIID and Vkappa family subgroup I, respectively. An expression vector of 125E11 single-chain Fv antibody fusion protein, in which VH and VL peptide were connected by a flexible linker (Gly(4)Ser)(3), was constructed. The scFv fusion protein was highly expressed in Escherichia coli mainly in inclusion body form. Using urea and pH gradient gel filtration method, the refolding of scFv was efficiently achieved. The refolding efficiency reached about 11% and 2.7 mg refolded scFv was obtained from 1L of culture. The binding activity and specificity of 125E11 scFv against PreS1(21-47)-containing antigen were also analyzed. SN - 1046-5928 UR - https://www.unboundmedicine.com/medline/citation/15866720/Gene_cloning_bacterial_expression_in_vitro_refolding_and_characterization_of_a_single_chain_Fv_antibody_against_PreS1_21_47__fragment_of_HBsAg_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S1046-5928(05)00047-1 DB - PRIME DP - Unbound Medicine ER -