Vitamin K and the biosynthesis of prothrombin. II. Structural comparison of normal and dicoumarol-induced bovine prothrombin.

Highly purified dicoumarol-induced bovine prothrombin, which does not bind calcium ions and has no prothrombin activity, has been structurally compared with normal prothrombin. Quantitative amino acid and carbohydrate analysis gave identical results for both prothrombins, as did analysis of the NHz-terminal and the COOH-terminal amino acids and molecular weight determination with the sodium dodecyl sulfate gel electrophoretic technique. Peptide maps of tryptic peptides prepared from the reduced and aminoethylated normal and dicoumarol-induced prothrombin were identical. These results suggest that the difference in properties between the two prothrombins are caused by a minor structural difference or a conformational difference. Ouchterlony immunodiffusion analysis gave a reaction of complete immunological identity between the two prothrombins, whereas the quantitative immunoprecipitation technique indicated antigenic difference between them. Furthermore, it was found that normal prothrombin has calcium ion-dependent antigenic determinants. The sedimentation coefficient, Stokes molecular radius, the titration curves for the tyrosine phenolic groups, and the fluorescence emission spectra were identical, which corroborates that the difference between the normal and the dicoumarol-induced prothrombin does not engage the entire molecule. The results obtained by polyacrylamide gel electrophoresis in 8 M urea may suggest that the difference includes an anomalous pairing of half-cystine residues.