Properties of FV and Fab fragments of the antibody McPC603 expressed in E. coli.
The FV and Fab fragments of the phosphorylcholine binding antibody McPC603 were functionally expressed in E. coli. This was achieved by the co-expression and co-secretion of both chains to the periplasm, where correct processing, folding and assembly occurred. Interestingly, the fraction of correctly folded Fab fragment is smaller than that of the Fv fragment in E. coli. The intrinsic hapten binding affinity was shown to be identical for the recombinant FV or Fab fragment, the whole antibody and the Fab fragment obtained by proteolysis from the mouse antibody. Fluorescence and crosslinking analyses showed that the FV fragment dissociates at high dilution, but that it is stabilized by hapten binding. The recombinant FV fragment was shown to have catalytic activity to hydrolyze choline-p-nitrophenyl carbonate and constitutes therefore a promising model system with which the structural requirements of catalytic antibodies can be studied by altering the protein itself.
Plückthun
A
A
Genzentrum der Universität München, Germany.
Glockshuber
R
R
Skerra
A
A
Stadmüller
J
J
D000818:Animals; D000906:Antibodies / Q000235:genetics / Q000276:immunology*; D004926:Escherichia coli / Q000235:genetics*; D006241:Haptens; D007140:Immunoglobulin Fab Fragments / Q000235:genetics / Q000276:immunology*; D007128:Immunoglobulin Fragments; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000276:immunology*; D007136:Immunoglobulins / Q000235:genetics / Q000276:immunology*; D007700:Kinetics; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology*
D000906:Antibodies; D006241:Haptens; D007140:Immunoglobulin Fab Fragments; D007128:Immunoglobulin Fragments; D007135:Immunoglobulin Variable Region; D007136:Immunoglobulins; C069242:McPC603 antibody; D011994:Recombinant Proteins; C055966:immunoglobulin Fv