Characterization of the binding properties of protein LG, an immunoglobulin-binding hybrid protein.
Protein LG is a 50-kDa hybrid molecule containing four Ig-light-chain-binding domains from protein L of Peptostreptococcus magnus and two IgG-Fe-binding repeats from streptococcal protein G. Here we analyse the binding of protein LG to Ig from several mammalian species. Protein LG was shown to bind human IgG of all subclasses and other Ig classes that carry kappa chains. The binding to human IgG was only marginally influenced by changes in temperature (4-37 degrees C) or salt concentration (0-1.6 M), and was stable over a wide pH range (pH 4-10). Protein LG bound to Ig from 11 of 12 mammalian species, including those of rabbit, mouse and rat. The affinity constants obtained for the interactions between protein LG and polyclonal IgG from rabbit (4.0 x 10(9) M-1), mouse (1.7 x 10(9) M-1) and rat (1.3 x 10(9) M-1) were similar to the value previously reported for the interaction between the hybrid protein and human polyclonal IgG (5.9 x 10(9) M-1). The interaction between protein LG and a mouse IgG mAb was not influenced by the presence of the specific protein antigen, nor was the binding of this antibody to its ligand affected by protein LG. Inhibition experiments demonstrated that the Ig-binding site of one of the fusion partners retained its ligand-binding capacity when the other component was occupied. Protein LG selectively absorbed 85-90% of the total Ig present in human and rabbit sera and 75-80% of the Ig in sera from mouse and rat. Human serum depleted of C1q, factor D and properdin and preabsorbed by protein LG could be used as a source for other complement factors. These data demonstrate that protein LG is a very versatile Ig-binding protein.
Kihlberg
B M
BM
Department of Cell and Molecular Biology, Section for Molecular Pathogenesis, Lund University, Sweden.
Sjöholm
A G
AG
Björck
L
L
Sjöbring
U
U
D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D001426:Bacterial Proteins / Q000378:metabolism*; D001667:Binding, Competitive; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007074:Immunoglobulin G / Q000378:metabolism; D007136:Immunoglobulins / Q000302:isolation & purification / Q000378:metabolism*; D007163:Immunosorbent Techniques; D066298:In Vitro Techniques; D051379:Mice; D010462:Peptostreptococcus / Q000378:metabolism; D011485:Protein Binding; D011817:Rabbits; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D013045:Species Specificity; D013291:Streptococcus / Q000378:metabolism
D000911:Antibodies, Monoclonal; D001426:Bacterial Proteins; C055047:Ig L-binding protein, Peptostreptococcus; C034284:IgG Fc-binding protein, Streptococcus; D007074:Immunoglobulin G; D007136:Immunoglobulins; D011994:Recombinant Proteins