Human coagulation factor IX. Isolation and characterization.
Human coagulation factor IX was purified by two ion-exchange chromatographies on DEAE-Sephadex A-50, heparin-Sepharose chromatography, hydroxyapatite chromatography and immunoadsorbent technique. Factor IX was homogeneous by ordinary and sodium dodecylsulphate disc electrophoresis, N-terminal amino acid analyses and ultracentrifugation and by immunological criteria. The following molecular data were observed: 1. Sedimentation equilibrium indicated a molecular weight of 66100 and sedimentation velocity gave S20,W = 3.97 S. A partial specific volume of -v = 0.712 ml/g was calculated from the amino acid and carbohydrate composition. 2. Sodium dodecylsulphate disc gel electrophoresis suggested a molecular weight of 65000. 3. Gel filtration indicated a Stokes radius of 4.08 nm, and 'a molecular weight' of 72000, as well as a diffusion coefficient D20,W = 5.15 X 10(-7) cm2 s-1 and a frictional ratio f/fo = 1.54. 4. Tyrosine was the N-terminal amino acid. The amino acid composition is described. Factor IX contained approximately 17.5% carbohydrate, which includes 4.7% hexose, 6.8% N-acetylhexosamine and 6% sialic acid. 5. Microheterogeneity of pure factor IX was demonstrated by isoelectric focusing. The isoelectric points of the major components lay within range of pH 4.0 to 4.6. 6. The antibody raised in rabbits against the pure factor IX did not react with the other vitamin-K-dependent coagulation factors measured by coagulation factor assays or immunodiffusion in gels.
D000596:Amino Acids / Q000032:analysis; D002241:Carbohydrates / Q000032:analysis; D002846:Chromatography, Affinity; D005164:Factor IX / Q000302:isolation & purification; D006801:Humans; D005779:Immunodiffusion; D007122:Immunoelectrophoresis; D007525:Isoelectric Focusing; D046911:Macromolecular Substances; D008970:Molecular Weight; D014443:Tyrosine / Q000032:analysis
D000596:Amino Acids; D002241:Carbohydrates; D046911:Macromolecular Substances; D014443:Tyrosine; D005164:Factor IX