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Tyrosine 69 of the first epidermal growth factor-like domain of human factor IX is essential for clotting activity
P E Hughes1, G Morgan, E K Rooney
1Sir William Dunn School of Pathology, University of Oxford, United Kingdom.
Epidermal growth factor (EGF)-like domains are present in many extracellular proteins including clotting factor IX. Those EGF-like domains with the consensus amino acid residues Asp/Asn, Asp/Asn, Asp*/Asn*, Tyr/Phe (where * denotes a beta-hydroxylated residue) bind calcium. Previous NMR studies of the isolated first EGF-like domain of human factor IX suggested that the first 3 consensus residues Asp-47, Asp-49, and Asp-64 were direct ligands for calcium. We now show from mutagenesis studies that Tyr-69 is not a direct ligand for calcium but is essential for the clotting activity of intact factor IX, specifically for the factor VIIIa-dependent activation of factor X. Surprisingly, Tyr-69 is not required for beta-hydroxylation of Asp-64.
Epidermal growth factor (EGF)-like domains are present in many extracellular proteins including clotting factor IX. Those EGF-like domains with the consensus amino acid residues Asp/Asn, Asp/Asn, Asp*/Asn*, Tyr/Phe (where * denotes a beta-hydroxylated residue) bind calcium. Previous NMR studies of the isolated first EGF-like domain of human factor IX suggested that the first 3 consensus residues Asp-47, Asp-49, and Asp-64 were direct ligands for calcium. We now show from mutagenesis studies that Tyr-69 is not a direct ligand for calcium but is essential for the clotting activity of intact factor IX, specifically for the factor VIIIa-dependent activation of factor X. Surprisingly, Tyr-69 is not required for beta-hydroxylation of Asp-64.