ABSTRACT
A three-dimensional (3D) graphic model of a single-chain Fv (scFv) which was derived from an anti-human placental acidic isoferritin (PAF) monoclonal antibody (Mab) was constructed by a homologous protein-predicting computer algorithm on Silicon graphic computer station.The structure, surface static electricity and hydrophobicity of scFv were investigated. Computer graphic modelling indicated that all regions of scFv including the linker, variable regions of the heavy (VH) and light (VL) chains were suitable. The VH region and the VL region were involved in composing the "hydrophobic pocket". The linker was drifted away VH and VL regions. The complementarity determining regions (CDRs) of VH and VL regions surrounded the "hydrophobic pocket". This study provides a theory basis for improving antibody affinity, investigating antibody structure and analyzing the functions of VH and VL regions in antibody activity.
ABSTRACT
A three-dimensional (3D) graphic model of a single-chain Fv (scFv) which was derived from an anti-human placental acidic isoferritin (PAF) monoclonal antibody (Mab) was constructed by a homologous protein-predicting computer algorithm on Silicon graphic computer station.The structure, surface static electricity and hydrophobicity of scFv were investigated. Computer graphic modelling indicated that all regions of scFv including the linker, variable regions of the heavy (VH) and light (VL) chains were suitable. The VH region and the VL region were involved in composing the "hydrophobic pocket". The linker was drifted away VH and VL regions. The complementarity determining regions (CDRs) of VH and VL regions surrounded the "hydrophobic pocket". This study provides a theory basis for improving antibody affinity, investigating antibody structure and analyzing the functions of VH and VL regions in antibody activity.
ABSTRACT
Objective:To obtain the functional single chain Fv antibody(scFv) against acidic isoferritin(AIF).Methods:An expression vector pPOW4c9 was constructed by subcloning AIF4c9 scFv gene into a heat-inducible bacterial expression plasmid pPOW3. Then recombinant vector was introduced into E.coli DH5? by electro-transformation. The soluble expression was performed by temperature induction. After purified by the Ni-chelating chromatography, the recombinant anti-AIF scFv was characterized.Results:Soluble expression of the scFv in E. coli was achieved. The yield of purified anti-AIF scFv was 1.6 mg/L. The recombinant protein recognized AIF specifically identified by ELISA and western blotting, and an affinity constant of scFv was 3.18?10~ -8 mol/L.Conclusion:The results indicate that recombinant soluble scFv retains the specific binding activity to AIF.