Single-chain Fv with Fc fragment of the human IgG1 tag: construction, Pichia pastoris expression and antigen binding characterization.
We report two expression vectors in Pichia pastoris that direct the synthesis of recombinant single chain antibody variable region (scFv), derived from anti-Z-DNA monoclonal antibody Z22. The first vector codes for a scFv fused to the Ig binding domain of staphylococcal Protein A. The second vector codes for the scFv fused to the Fc fragment of the human IgG1. The fusion partner simplified the detection and purification of the secreted protein. These constructs yielded high level expression of an scFv with specific binding activity toward a Z form of DNA, with binding activity comparable to that of the scFv molecule produced in an Escherichia coli expression system and the original monoclonal antibody.
Andrade
E V
EV
Laboratório de Biologia Molecular, Universidade de Brasília, Brasília, Brasil.
Albuquerque
F C
FC
Moraes
L M
LM
Brígido
M M
MM
Santos-Silva
M A
MA
D000941:Antigens / Q000276:immunology*; D001483:Base Sequence; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005822:Genetic Vectors; D006801:Humans; D007074:Immunoglobulin G / Q000235:genetics*; D007135:Immunoglobulin Variable Region / Q000235:genetics* / Q000276:immunology; D009838:Oligodeoxyribonucleotides; D010843:Pichia / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification
D000941:Antigens; D007074:Immunoglobulin G; D007135:Immunoglobulin Variable Region; D009838:Oligodeoxyribonucleotides; D011993:Recombinant Fusion Proteins