Localization within the 106 N-terminal amino acids of von Willebrand factor (vWF) of the epitope corresponding to a monoclonal antibody which inhibits vWF binding to factor VIII.
We have used an in vitro transcription-translation system to localize the epitope corresponding to a monoclonal antibody (MAb 418) to vWF which specifically inhibits its binding to F. VIII. We have subcloned 1.5 Kb of vWF cDNA encoding for the N-terminal part of the vWF subunit which contains a binding site for F.VIII into the Bluescribe expression vector. After in vitro transcription and translation using the resulting construction (pBS-TG3522), a polypeptide of mol wt 60,000 (AA 1-495) was immunoprecipitated with a polyclonal antibody to vWF and with MAb 418. The MAb 418 epitope was further localized by reducing the size of the cDNA insert. This resulted in the production of two polypeptides of 18,000 (AA 1 to 142) and 13,000 (AA 1 to 106) which both retained reactivity with MAb 418. Thus the epitope corresponding to MAb 418 is localized to the first 106 AA of the mature vWF subunit.
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INSERM U. 143, Hôpital de Bicêtre, le Kremlin Bicêtre, France.
Ribba
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Meulien
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Meyer
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D000911:Antibodies, Monoclonal / Q000276:immunology*; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes; D005169:Factor VIII / Q000378:metabolism*; D006801:Humans; D008970:Molecular Weight; D010446:Peptide Fragments / Q000276:immunology; D011485:Protein Binding; D011994:Recombinant Proteins / Q000276:immunology; D014841:von Willebrand Factor / Q000276:immunology* / Q000378:metabolism / Q000648:ultrastructure
D000911:Antibodies, Monoclonal; D000939:Epitopes; D010446:Peptide Fragments; D011994:Recombinant Proteins; D014841:von Willebrand Factor; D005169:Factor VIII