Isolation and characterization of human polyreactive pneumococcal polysaccharide antibodies

Abstract

Natural antibodies serve as the body's first line of defense against pneumococcal challenge. Polyreactive human pneumococcal polysaccha-ride IgG antibodies have not been extensively studied. We analyzed human polyreactive anti-bodies that bind multiple pneumococcal poly-saccharides, including PPS14 and PPS23F. These antibodies were isolated from single pneumococcal polysaccharide specific B cells allowing for the analysis of human immu-noglobulins with natively paired variable regions. Although isolated individually, these antibodies demonstrated similar characteristics. Most an-tibodies possessed a variable light chain with a CDR3 length made up of nine amino acids and relatively high number of flexible amino acids in combined VH/VL. While these antibodies were polyreactive and structurally alike, kinetic analy-sis revealed unique K D values. Variable chains are responsible for antigen recognition whereas antibody fine specificity is affected by isotype structure. To investigate the contribution of the constant region of these isotypes and their ef-fect on antibody avidity to pneumococcal poly-saccharide, the polyreactive variable regions were expressed as IgG1 or IgG2 and subjected to kinetic analysis. The IgG1 antibodies uni-formly had a stronger avidity to PPS14 and PPS23F compared to IgG2. To further document the importance of the constant region in anti-body avidity and fine specificity, analysis of an-tibody F(ab)'2 fragment binding to PPS14 and PPS23F resulted in similar K D values. These studies suggest that antigen recognition by polyreactive antibodies is determined by a con-served variable light chain CDR3 length and longer, more flexible variable heavy CDR3s when compared to pneumococcal polysaccha-ride-specific sequences while differences in specific avidities are modulated by antibody isotype.