The complete amino acid sequence of hen ovalbumin, comprising 385 residues, has been determined. The sequence was deduced from the 17 cyanogen bromide fragments and from peptides derived by digestion with a number of proteolytic enzymes. The molecular weight of the polypeptide chain of ovalbumin is 42699.Ovalbumin has four sites of postsynthetic modification ; in addition to the acetylated N terminus, the carbohydrate moiety is located at Asn-292, and the two phosphorylated serines are at residues 68 and 344. The 'signal sequence' of ovalbumin is between residues 234 and 252. The heptapeptide released during the conversion of ovalbumin to plakalbumin by subtilisin digestion corresponds to residues 346 -352. The hen ovalbumin polymorphism characterised by an Asn-tAsp replacement results from a mutation at residue 31 1.The amino acid sequence of ovalbumin deduced from these amino acid sequence studies is in complete agreement with the sequence of mRNA determined by McReynolds et al. [Nature (Lond.) 273, 723 -728 (1978)].Ovalbumin is the most abundant protein in egg white, and is readily purified and crystallised in gram quantities [l].

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Mg2+ anticoagulated PPP (20mM MgCl2) was applied at RT to a Zn2+ immobilised biscarboxymethylamino Sepharose 4B column and eluted with 20mM Tris, 20mM MgCl2, 2.5mM CaCl2 0.15M NaCl buffer pH 7.4. Following collection of the wash-through fractions, bound proteins were developed through application of linear (0 to 35mM) imidazole gradients.All fractions were screened for F.II, V, IX, X, vWF:Ag, Protein C, Fg, Fn and 2-macroglobulin by ELISA, VIII:Ag by IRMA, and VIII:C by both 1-stage and 2-stage bioassay methods. VIIIrAg (60-90% yield), vWF:Ag (100%), VIII:C 1-stage activity (35%), Fn (> 60%), Fg (> 80%), V (50%) and a2-macroglobulin (> 70%) were located only in the gradient fractions. Two distinct peaks demonstrated shortening of the 2-stage VIII:C assay:- one co-eluting with the 1-stage VIII:C activity and another major peak in the wash-through fractions where antigenic determinants of F.II, IX, X and part of the protein C were located and partially resolved from each other. Only F.II.-Ag co-eluted with the “ 2-stage VIII:C” activity. Similar observations were found in Mg2+ -anticoagulated severe Haemophilia A plasma and in citrated PPP developed with 20mM MgCl2, 20mM Tris buffer. A1(0H)3 treatment abolished the activity from citrate -, but not from Mg2+ -anticoagulated plasma.The relevant fractions showed no activities in F.V, VII, VIII:C, IX or X 1-stage bioassays. They did not clot Fg and did not contain detectable Xa or Ila as assessed by S-2222, S-2238 or S-2288. Following incubation with specific antisera against IgG, II, V, VII:Ag, X, protein C, a- and g- lipoproteins, and plasminogen only anti-II inhibited this "2-stage VIII:C" activity. 2-stage VIII:C assays depend upon Ca2+ -dependent generation of Xa. Since the activity in the wash-through fractions could not be ascribed to VIII, Xa, or Ila the results would indicate the presence of a hitherto undescribed Factor X activator activity.