Citraconyl-BSA by Trypsin

75 column. Two molecules of antibody were bound per mole of peptide. Intactness of the disulfide bonds was essential for maintenance of the inhibitory activity of the peptide. Its immunochemical reactivity was completely destroyed on rupture of the disulfide bonds by performic acid oxidation or by reduction followed by carboxymethylation. Also, cleavage at lysyl peptide bonds, without rupture of the disulfide bonds, destroyed the inhibitory activity of the peptide, because of scission of the polypeptide chain at (or in proximity of) the antigenic reactive sites. Since the peptide comprised less than a third of BSA molecule but accounted for almost all the BSA antigenic reactivity, it was concluded that native BSA carried equivalent antigenic reactive sites (Atassi et aL, 1976 ). 

A further support to this view has been afforded from our isolation of another fragment from a tryptic digest of bovine serum albumin (Habeeb and Atassi, 1976a,6). The fragmentation pattern of crystalline native BSA by trypsin was studied (Habeeb and Atassi, 1976a,6) in aqueous solution under various conditions with regard to the yield and size of the 

BSA (Atassi et al., 19766), the immunochemical results can be explained (Atassi et al., 19766 Habeeb and Atassi, 1976a,6) only by the concept that BSA carries repeating and inununochemically equivalent or simUar antigenic reactive sites. 

Protein or fragment Antiserum B5 Antiserum B7 Antiserum 487 c Antiserum 488 Antiserum 489  

Reactions of adsorbed antiserum relative to unadsorbed antiserum  

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However, this assignment can be considered as tentative since the amino acid sequence of BSA was not available at that time. This mode of fragmentation of citraconylated BSA by chymotrypsin is in contrast to the fragmentation of citraconyl-BSA by trypsin (Atassi et aL, 1976 ), in which the main fragment corresponded to sequence Phen-Argigg (derived from the amino-terminal region). 

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