Homology modelling and bivalent single-chain Fv construction of anti-HepG2 single-chain immunoglobulin Fv fragments from a phage display library.

Abstract	We prepared single-chain immunoglobulin Fv fragments (scFv) SLH10 specific for the HepG2 cell line after biopanning from a large human-naive phage display library (Griffin. 1 Library). The three-dimensional (3D) structure of SLH10 was modelled by the Insight II molecule simulation software.The structure was refined using the molecular dynamics method.The structures with the least steric clashes and lowest energy were determined finally. The optimized structures of heavy (VH) and light (VL) variable chains of SLH10 scFv were obtained.Then SLH10 bivalent single-chain Fv (BsFv) was constructed that would be suitable for high-affinity targeting.SLH10 BsFv was generated by linking scFvs together and identified by sequencing. Its expression products were confirmed by western blot analysis.The relative molecular masses of scFv and BsFv were approximately 30 kDa and 60 kDa,respectively. Flow cytometry revealed that SLH10 BsFv bound the selected cell lines with greater signal intensity than the parental scFv. The improved antigen binding of SLH10 BsFv may be useful for immunodiagnostics or targeted gene therapy for liver cancer.
Authors	Ming Ni, Bing Yu, Yu Huang, Zhenjie Tang, Ping Lei, Xin Shen, Wei Xin, Huifen Zhu, Guanxin Shen
Journal	Journal of biosciences (J Biosci) Vol. 33 Issue 5 Pg. 691-7 (Dec 2008) ISSN: 0250-5991 [Print] India
PMID	19179757 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
Chemical References	Antibodies Immunoglobulin Fragments Immunoglobulin Heavy Chains Immunoglobulin Light Chains Peptide Library immunoglobulin Fv
Topics	Antibodies Cell Line Hepatocytes (immunology) Humans Immunoglobulin Fragments (chemistry, immunology) Immunoglobulin Heavy Chains Immunoglobulin Light Chains Models, Biological Models, Molecular Peptide Library Protein Binding Protein Conformation Sequence Homology, Amino Acid

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