Generation and characterization of peptide-specific antibodies that inhibit von Willebrand factor binding to glycoprotein IIb-IIIa without interacting with other adhesive molecules. Selectivity is conferred by Pro1743 and other amino acid residues adjacent to the sequence Arg1744-Gly1745-Asp1746

Academic Article
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publication date

  • 1988

abstract

  • We have generated antibodies against a synthetic peptide corresponding to the sequence of human von Willebrand factor (vWF) between residues Glu1737-Ser1750 which includes the Arg-Gly-Asp sequence common to several adhesive molecules. Two anti-peptide antibodies, one polyclonal, and one monoclonal reacted with native vWF and inhibited its binding to platelet glycoprotein (GP) IIb-IIIa, but showed negligible cross-reactivity with fibrinogen, fibronectin, and vitronectin, three other molecules that contain the sequence Arg-Gly-Asp and bind to platelets. The structural bases for the specificity of the two antibodies were evaluated by testing the ability of peptides homologous to the parent sequence, but with single amino acid substitutions, to neutralize the binding of the two antibodies to vWF. The substitution of Pro1743, the residue immediately adjacent to the Arg-Gly-Asp sequence on the amino-terminal side, with Phe resulted in a peptide that failed to interact with either antibody. Thus, Pro1743 is important for maintaining a peptide conformation recognized by two antibodies specific for the GP IIb-IIIa-binding domain of vWF. Other residues important for optimal peptide reactivity with the polyclonal antibody were Ser1742, Arg1744, and Gly1745, whereas Gly1741, Gly1745, and Asp1746, but not Arg1744, were important for reactivity with the monoclonal antibody. The epitopes of both antibodies, therefore, included at least 2 of the residues in the sequence Arg-Gly-Asp considered the common cell-binding site of adhesive molecules that interact with GP IIb-IIIa. Nevertheless, both antibodies reacted only with vWF. These studies demonstrate that peptide-specific antibodies, unlike the promiscuous GP IIb-IIIa receptor, can recognize distinctive structural characteristics of the cell-binding domain of adhesive molecules imposed by residues adjacent to the sequence Arg-Gly-Asp.