Purification and characterization of dicoumarol-induced prothrombins. Evidence of 5-Gla variant with two (or more)...

A 5-Gla prothrombin, adsorbable onto barium oxalate and containing an admixture of one- and two-chain molecules, has been purified from the plasma of steers on a dicoumarol regimen for three years. Double-chain molecules were not detected in any of our six other variants containing zero to nine Gla residues. In this double-chain variant, the peptide bond between Arg52-Asn appears to be missing, as the preparation showed amino-terminal alanine and aspartic acid, and released both double- and single-chain F1 upon digestion with thrombin. The molecular masses of the F1 and its heavy chain were 24,000 and 19,000 daltons, respectively. The double-chain variant was similar to its single-chain counterpart [Malhotra, O.P. (1979) Thromb. Res. 14, 439-448] in that it (a) yielded thrombin equivalent to 30% of normal in three hr, (b) moved in the alpha 1 region in the absence of calcium ions and between the alpha 1 and alpha 2 macroglobulins in the presence of calcium ions, (c) contained five gamma-carboxyglutamyl residues (Gla), and (d) showed comparable antigenic activity with normal prothrombin.