Abstract
Pursuing an autoimmune model for the etiology of poststreptococcal glomerulonephritis, protein antigens isolated from the cytoplasmic membrane of nephritogenic group A Type 12 Streptococcus pyogenes were immunochemically characterized using antistreptococcal cell membrane (SCM) monoclonal antibody (MAb) cross-reactive with glomerular basement membrane (GBM). Low molecular weight (9.2, 7.0, 4.7, 2.3 kDa) HPLC-purified SCM polypeptide antigens were characterized by competitive inhibition and equilibrium dialysis. Competitive inhibition of the MAb by different sized SCM polypeptide antigens showed an inverse relationship between the size of these antigens and the molar amount required to obtain 50% inhibition of the MAb, confirming previous observations that suggested that these SCM antigens exhibit increasing epitope concentration with increasing size, that is constant epitope density. The observed changes in epitope concentration correlated with differences in the valence and affinity of the MAb as determined by equilibrium dialysis. The Kds of the MAb for 9.2-, 7.0-, 4.7-, and 2.3-kDa SCM antigens ranged from 7.42 x 10-7 to 1.15 x 10-5. The experimentally determined MAb valence for these antigens was 2 for the 9.2-kDa antigen and approached 10 for the smaller antigens. Finally, the similarity of these SCM antigens was reflected in similar amino acid compositions; of note, these data agreed with the compositions previously reported for sized GBM antigens. Concentrations of Asp, Thr, Ser, Glu, Gly, Ala, Val, Ile, and Leu paralleled increasing epitope concentration. Apparent N-terminal blocking prevented sequencing of these peptides, but these immunochemical data suggest that intact SCM antigen recognized by the anti-SCM MAb consists of repeating epitopes, an observation consistent with the cytoplasmic membrane source of the antigen.
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