Vitamin K and the Biosynthesis of Prothrombin

Abstract The 25,000 Mr fragment obtained by thrombin digestion of normal prothrombin has been further compared with the corresponding fragment from dicoumarol-induced prothrombin. In contrast with the fragment from normal prothrombin, the one from dicoumarol-induced prothrombin does not bind calcium (Stenflo, J. (1973) J. Biol. Chem. 248, 6325). After cyanogen bromide degradation of this material, a fragment with approximately 75 amino acid residues constituting the NH2-terminal part of prothrombin was isolated by gel filtration. The fragment from normal prothrombin bound calcium, while the fragment from the dicoumarolinduced prothrombin did not. The fragments from the two prothrombins did not contain N-acetylglucosamine and had identical amino acid compositions. From tryptic digests of the completely reduced and S-carboxymethylated fragment from normal prothrombin two peptides were isolated which differed from the corresponding peptides in digests of the abnormal prothrombin. One peptide from normal prothrombin, which contained residues 4 to 10, had an anodal electrophoretic mobility too high to be consistent with the amino acid composition, thereby indicating a prosthetic group with a charge of at least -3. The corresponding peptide from abnormal prothrombin had an electrophoretic mobility consistent with the amino acid composition. The other peptide in normal prothrombin contained residues 12 to approximately 34. It had arginine residues inaccessible to trypsin. In abnormal prothrombin, this peptide was replaced by smaller peptides. Both peptides from normal prothrombin bound calcium. The properties of the two peptides from normal prothrombin, together with the comparison with the corresponding peptides from abnormal prothrombin, provide strong evidence for the presence of prosthetic groups in the peptides from normal prothrombin. Since there is no evidence of prosthetic groups in the peptides from dicoumarol-induced prothrombin these must have been attached to the prothrombin polypeptide chain by the action of vitamin K.