The alternative binding site for protein A in the Fab fragment of immunoglobulins.
Twenty-six new human or murine monoclonal immunoglobulins (IgM, IgA, murine IgG1 or human IgG3) with a known V-region sequence were tested for alternative (non-Fc) binding to Staphylococcal protein A. Seven of them did not bind at all. Four immunoglobulins (all mouse IgG1) were bound but easily eluted (at pH 6). They were probably bound via the Fc part. All eleven were classified as negative for alternative binding. Fifteen immunoglobulins were found to bind more firmly; they came off the protein A column at pH 4-3 (alternative binders). Amino acid sequences of immunoglobulins that have been typed in the present work or earlier (25 binders and 26 non-binders) were compared. The light chain, the C region of the heavy chain and the D and JH segments look irrelevant for alternative binding. The N-terminal portion (amino acids 1-94) of the H chain probably forms the ligand of protein A. A peptide making the ligand cannot be reliably localized within this stretch but binder proteins had a high homology in residues 6-29. All mouse immunoglobulins expressing VH genes of families J606 or S107 were alternative binders; those expressing other families were non-binders.
Ibrahim
S
S
Department of Bacteriology and Immunology, University of Helsinki, Finland.
Kaartinen
M
M
Seppälä
I
I
Matoso-Ferreira
A
A
Mäkelä
O
O
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry; D001483:Base Sequence; D001665:Binding Sites; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000378:metabolism*; D051379:Mice; D008806:Mice, Inbred AKR; D008807:Mice, Inbred BALB C; D008808:Mice, Inbred CBA; D008815:Mice, Inbred Strains; D008969:Molecular Sequence Data; D013205:Staphylococcal Protein A / Q000737:chemistry* / Q000378:metabolism
D000911:Antibodies, Monoclonal; D007140:Immunoglobulin Fab Fragments; D013205:Staphylococcal Protein A