scFv multimers of the anti-neuraminidase antibody NC10: length of the linker between VH and VL domains dictates precisely the transition between diabodies and triabodies.
Single-chain Fv antibody fragments (scFvs) incorporate a polypeptide linker to tether the VH and VL domains together. An scFv molecule with a linker 5-12 residues long cannot fold into a functional Fv domain and instead associates with a second scFv molecule to form a bivalent dimer (diabody). Direct ligation of VH and VL domains further restricts association and forces three scFv molecules to associate into a trivalent trimer (triabody). We have defined the effect of linker length on scFv association by constructing a series of scFvs from anti-neuraminidase antibody NC10 in which the linker varied from one to four glycine residues. NC10 scFv molecules containing linkers of three and four residues showed a strong preference for dimer formation (diabodies), whereas a linker length of one or two glycine residues prevented the formation of diabodies and directed scFv association into trimers (triabodies). The data suggest a relatively strict transition from dimer (diabody) to trimer (triabody) upon reduction of the linker length from three to two glycine residues. Modelling studies are consistent with three residues as the minimum linker length compatible with diabody formation. Electron microscope images of complexes formed between the NC10 scFv multimers and an anti-idiotype Fab' showed that the dimer was bivalent for antigen binding and the trimer was trivalent.
Atwell
J L
JL
CSIRO Molecular Science and CRC for Diagnostic Technologies, 343 Royal Parade, Parkville, Victoria, Australia. john.atwell@molsci.csiro.au
Breheney
K A
KA
Lawrence
L J
LJ
McCoy
A J
AJ
Kortt
A A
AA
Hudson
P J
PJ
D000906:Antibodies / Q000737:chemistry*; D019281:Dimerization; D005998:Glycine / Q000737:chemistry; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry; D007128:Immunoglobulin Fragments / Q000737:chemistry*; D007143:Immunoglobulin Heavy Chains / Q000737:chemistry*; D007130:Immunoglobulin Idiotypes / Q000737:chemistry; D007147:Immunoglobulin Light Chains / Q000737:chemistry*; D008854:Microscopy, Electron; D008958:Models, Molecular; D009439:Neuraminidase / Q000276:immunology*; D011487:Protein Conformation
D000906:Antibodies; D007140:Immunoglobulin Fab Fragments; D007128:Immunoglobulin Fragments; D007143:Immunoglobulin Heavy Chains; D007130:Immunoglobulin Idiotypes; D007147:Immunoglobulin Light Chains; C055966:immunoglobulin Fv; D009439:Neuraminidase; D005998:Glycine