Mammalian cell expression of dimeric small immune proteins (SIP).
We have designed and expressed bivalent small immune proteins (SIP) based on scFv fragments connected through a short linker of four amino acids to the CH3 domain of the human immunoglobulin gamma 1 H-chain. Three different versions have been designed and expressed in mammalian cells. In one construct a cysteine residue was included in the last amino acid of the flexible 15-amino acid long linker connecting the V(L) and V(H) domains, thus creating a disulphide bond stabilized molecule. A version with a shorter (five amino acids) V(L)/V(H) linker was also produced and shown to be efficiently assembled and secreted. All three SIPs form dimers retaining their antigenic specificity in Western blotting and having a comparable functional affinity (avidity) as determined by ELISA.
Li
E
E
International Centre for Genetic Engineering and Biotechnology, Area Science Park, Trieste, Italy.
Pedraza
A
A
Bestagno
M
M
Mancardi
S
S
Sanchez
R
R
Burrone
O
O
D000818:Animals; D015153:Blotting, Western; D016466:CHO Cells; D006224:Cricetinae; D019281:Dimerization; D004797:Enzyme-Linked Immunosorbent Assay; D005822:Genetic Vectors / Q000138:chemical synthesis; D006801:Humans; D007128:Immunoglobulin Fragments / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics*; D007135:Immunoglobulin Variable Region / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics*; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D014176:Protein Biosynthesis; D011506:Proteins / Q000235:genetics* / Q000276:immunology*; D014407:Tumor Cells, Cultured
D007128:Immunoglobulin Fragments; D007135:Immunoglobulin Variable Region; D011506:Proteins