Immunochemical studies of human fibrinopeptide A using synthetic peptide homologues

Abstract

Previous studies indicated that rabbit antisera R2 and R33 to human fibrinopeptide A differ markedly in terms of cross-reactivity with fibrinogen and fibrinopeptide A-containing fragments of the fibrinogen molecule. Antiserum specificity was characterized by comparison of inhibition of binding to radiolabeled tyrosyl fibrinopeptide A (produced by synthetic fragments and enzymatic digests of the fibrinopeptide A molecule) vs. the complete fibrinopeptide sequence (A.alpha. 1-16). Synthetic COOH-terminal homologues through the dodecapeptide (A.alpha. 5-16) exhibited less than 16% immunoreactivity with R33 antiserum, which cross-reacts extensively with fibrinogen and fibrinopeptide A-containing fibrinogen fragments. The synthetic COOH-terminal decapeptide (A.alpha. 7-16) gave 100% immunoreactivity with R2 antiserum, which cross-reacts minimally with fibrinogen and fibrinopeptide A-containing fibrinogen fragments. Synthetic homologues smaller than A.alpha. 7-16, such as A.alpha. 9-16 and A.alpha. 7-11, reacted only minimally with R2 antiserum. Carboxypeptidase B digests of fibrinopeptide A retained less than 25% of their initial immunoreactivity with R2 antiserum. The antigenic determinants of R2 immunoreactivity reside entirely within the COOH-terminal 10-residue sequence of fibrinopeptide A. Phe-8, Asp-7 and Arg-16 contribute significantly to R2 immunoreactivity. The R2 antigenic determinants appear to be significantly less accessible to reaction with antibody than the R33 determinants when the fibrinopeptide is attached to its parent .alpha. chain. A possible mechanism for the sequestration is discussed.