Insilico analysis of three different tag polypeptides with dual roles in scFv antibodies.
Single chain fragment variable (scFv) antibodies are composed of variable heavy (VH) and variable light (VL) domains that are joined by a polypeptide linker. Typically, [(Gly4Ser) n] sequence is used as a linker to retain the integrity of the antigen-binding domain. Due to its low immunogenicity, this sequence cannot be used as a tag for scFv detection and purification. Several evidences have shown that the addition of an N or C-terminal tag for scFv detection and purification will result in the decreased expression and binding capacity of this antibody fragment. In this study, we substituted the traditional linker (GGGGS) with His-tag, C-myc or E-tag sequences through molecular modeling. Stability and integrity of all models were assessed by molecular dynamic (MD) simulation. Based on MD simulation analysis, the model containing E-tag sequence as a linker indicated more stability compared to other molecules. The results suggest that E-tag not only can be substituted for the traditional linker, also eliminates the necessity of using additional tag for scFv detection and purification.
Faculty of Advanced Medical Sciences and Technologies, Shiraz University of Medical Sciences, Shiraz, Iran; Recombinant antibody laboratory, Department of immunology, Shiraz University of Medical Sciences, Shiraz, Iran.
Mozafar
M
Mohammadi
Recombinant antibody laboratory, Department of immunology, Shiraz University of Medical Sciences, Shiraz, Iran; Shiraz HIV/AIDS research center, Shiraz University of Medical Sciences, Shiraz, Iran. Electronic address: nejatollaf@sums.ac.ir.
Foroogh
F
Nejatollahi
School of Pharmacy, Shiraz University of Medical Sciences, Shiraz, Iran.
Amirhossein
A
Sakhteman
Department of Biotechnology, School of Veterinary Medicine, Shiraz University, Shiraz, Iran.
Neda
N
Zarei
D000595:Amino Acid Sequence; D003198:Computer Simulation; D006860:Hydrogen Bonding; D056004:Molecular Dynamics Simulation; D010455:Peptides / Q000737:chemistry*; D017434:Protein Structure, Tertiary; D015203:Reproducibility of Results; D016415:Sequence Alignment; D057127:Single-Chain Antibodies / Q000737:chemistry*; D040681:Structural Homology, Protein
D010455:Peptides; D057127:Single-Chain Antibodies
MD simulation; Molecular modeling; Polypeptide linker; ScFv stability