Comparison of amino acid sequence of bovine coagulation Factor IX (Christmas Factor) with that of other vitamin K-dependent plasma proteins.

Abstract

The amino acid sequence of bovine blood coagulation Factor IX (Christmas Factor) is presented and compared with the sequences of other vitamin K-dependent plasma proteins and pancreatic trypsinogen. The 416-residue sequence of Factor IX was determined largely by automated Edman degradation of two large segments, containing 181 and 235 residues, isolated after activating Factor IX with a protease from Russell's viper venom. Subfragments of the two segments were produced by enzymatic digestion and by chemical cleavage of methionyl, tryptophyl, and asparaginyl-glycyl bonds.Comparison of the amino acid sequences of Factor IX, Factor X, and Protein C demonstrates that they are homologous throughout. Their homology with prothrombin, however, is restricted to the amino-terminal region, which is rich in y-carboxyglutamic acid, and the carboxyl-terminal region, which represents the catalytic domain of these proteins and corresponds to that of pancreatic serine proteases. Factor IX (Christmas Factor) is the zymogen of a serine protease that participates in the middle phase of the intrinsic pathway of blood coagulation (1). Like several other plasma proteinsi.e., prothrombin, Factor X, Factor VII, Protein C, and Protein S-Factor IX requires vitamin K for its biosynthesis (2-4). Individuals lacking Factor IX (Christmas disease or hemophilia B) show bleeding symptoms essentially identical to those of classic hemophilia or hemophilia A (Factor VIII deficiency) (5, 6). The activity of Factor IX is also depressed in the plasma of patients treated with coumarin analogs, but in such cases the effect can be reversed by the administration of vitamin K (6).Bovine and human Factors IX have been purified and characterized (7)(8)(9)(10)(11)(12) Previous comparison of the partial sequence of bovine Factor IX with the sequences of bovine prothrombin and Factor X revealed homologous regions, indicating that these three proteins may have evolved from a common ancestral protein (16,17). Recently, the light and heavy chains of bovine Protein C, another vitamin K-dependent plasma protein, have been found to be homologous with Factor X (18,19). In the present communication, the complete sequence of Factor IX is presented.Comparison with the sequences of Factor X, Protein C, and prothrombin reveals areas of homology as well as regions lacking obvious similarity. The experimental details of the sequence determination will be published elsewhere. METHODS Factor IX, isolated from bovine plasma (7) and converted to Factor IXaa by a protease from Russell's viper venom (15), was obtained through the courtesy of K. Fujikawa. Factor IXaa was reduced with dithioerythritol and S-alkylated with 4-vinylpyridine. The two peptide chains (segment N and segment C) were separated on a column of SP-Sephadex C-25 by applying a linear gradient of sodium formate buffers in the presence of 7 M urea (20).Segment N (residues 1-181) was divided into two fragments (CB I and CB II), by cleavage with cyanogen bromide under conditions that avoid cleavage at y-carbo...

Isolation and N‐terminal amino acid sequence of protein Z, A γ‐carboxyglutamic acid containing protein from bovine plasma

“…Samples from the column effluent were subjected to alkaline hydrolysis and Gla was found mainly in a fraction eluting at 0.75 M NH4HC0s. The amino acid composition of pooled material from this fraction is shown in table 1, and is very similar to those of the Gla-domains of prothrombin [7,8], factor IX [9], factor X [lO,ll], and protein C [ 121. Further evidence that protein Z has a Glaregion of similar structure was obtained from N-terminal sequence analysis of protein Z.…”

“…Alignment of the N-terminal sequences of the known Gla-proteins from bovine plasma. Prothrombin [7], factor X [lO;ll], factor IX [9], factor VII [22], protein C [2], and protein S [4].…”

“…This paper describes the purification, amino acid composition, and N-terminal amino acid sequence of protein Z. Furthermore, a peptide with an amino acid composition similar to those of the N-terminal Gla-domains (residues -5-42) of prothrombin [7,8], factor IX [9], factor X [lO,ll], and protein C [12] has been isolated after digestion of protein Z with chymotrypsin.…”

Localization of the specific binding site for magnesium(II) ions in factor IX

“…Changes in amino acid residues should induce changes in tertiary structure. It is possible that the size of metal-binding pocket formed by these residues is reduced such that a Ca 2+ ion can no longer gain [20] and human [21] factor IX, human factor X [22], human factor VII [23], human protein C (PC) [24] and human prothrombin (PT) [25] are aligned. Residues identical to those in both human and bovine factor IX are shaded.…”

Binding of the Factor IX γ-Carboxyglutamic Acid Domain to the Vitamin K-dependent γ-Glutamyl Carboxylase Active Site Induces an Allosteric Effect That May Ensure Processive Carboxylation and Regulate the Release of Carboxylated Product

“…It catalyzes the post-translational modification of specific glutamic acid residues to ␥-carboxylglutamic acid in a number of vitamin K-dependent proteins. In these vitamin K-dependent proteins, multiple glutamic acid residues in the amino-terminal Gla 1 domain are modified (1)(2)(3). The existence of these multiple ␥-carboxylglutamic acid residues allows the Gla domain to form the calcium-dependent conformation required for the activity of these vitamin K-dependent proteins (4,5).…”