Blood clotting factor IX B(M) Nagoya. Substitution of arginine 180 by tryptophan and its activation by α-chymotrypsin and rat mast cell chymase

K. Suehiro, Shun-Ichiro Kawabata, T. Miyata, H. Takeya, J. Takamatsu, K. Ogata, T. Kamiya, H. Saito, Y. Niho, S. Iwanaga

研究成果: ジャーナルへの寄稿記事

17 引用 (Scopus)

抄録

Factor IX B(M) Nagoya (IX Nagoya) is a natural mutant of factor IX responsible for severe hemophilia B. A patient with this mutant is characterized by a markedly prolonged ox brain prothrombin time. IX Nagoya was purified from the patient's plasma by immunoaffinity chromatography with an anti-factor IX monoclonal antibody column. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that treatment of IX Nagoya with factor XIa/Ca2+ resulted in cleavage only at the Arg145-Ala146 bond. Reversed-phase high performance liquid chromatography of a trypsin digest of IX Nagoya showed an aberrant peptide, which was further digested with proteinase Asp-N. Primary structure analysis of one of the Asp-N peptides revealed that Arg180 is replaced by Trp. An essentially complete (99%) amino acid sequence of IX Nagoya was obtained by sequencing fragments derived from a lysyl endopeptidase digest in which no other substitutions in the catalytic triad or substrate binding site were found. We also found that IX Nagoya is activated by α-chymotrypsin or rat mast cell chymase by monitoring the rate of factor X activation using a fluorogenic peptide substrate in the presence of factor VII, phospholipids, and Ca2+. These results indicate that the substitution of Arg180 by Trp impairs the cleavage by factor XIa required for activation of this zymogen and that the substitution causes hemophilia B(M).

元の言語英語
ページ(範囲)21257-21265
ページ数9
ジャーナルJournal of Biological Chemistry
264
発行部数35
出版物ステータス出版済み - 12 1 1989

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Chymases
Factor IX
Blood Coagulation Factors
Chymotrypsin
Blood Coagulation
Mast Cells
Tryptophan
Arginine
Rats
Blood
Substitution reactions
Chemical activation
Factor XIa
Hemophilia B
lysyl endopeptidase
Peptides
Factor X
Enzyme Precursors
Factor VII
Prothrombin Time

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

これを引用

Blood clotting factor IX B(M) Nagoya. Substitution of arginine 180 by tryptophan and its activation by α-chymotrypsin and rat mast cell chymase. / Suehiro, K.; Kawabata, Shun-Ichiro; Miyata, T.; Takeya, H.; Takamatsu, J.; Ogata, K.; Kamiya, T.; Saito, H.; Niho, Y.; Iwanaga, S.

:: Journal of Biological Chemistry, 巻 264, 番号 35, 01.12.1989, p. 21257-21265.

研究成果: ジャーナルへの寄稿記事

Suehiro, K, Kawabata, S-I, Miyata, T, Takeya, H, Takamatsu, J, Ogata, K, Kamiya, T, Saito, H, Niho, Y & Iwanaga, S 1989, 'Blood clotting factor IX B(M) Nagoya. Substitution of arginine 180 by tryptophan and its activation by α-chymotrypsin and rat mast cell chymase', Journal of Biological Chemistry, 巻. 264, 番号 35, pp. 21257-21265.
Suehiro, K. ; Kawabata, Shun-Ichiro ; Miyata, T. ; Takeya, H. ; Takamatsu, J. ; Ogata, K. ; Kamiya, T. ; Saito, H. ; Niho, Y. ; Iwanaga, S. / Blood clotting factor IX B(M) Nagoya. Substitution of arginine 180 by tryptophan and its activation by α-chymotrypsin and rat mast cell chymase. :: Journal of Biological Chemistry. 1989 ; 巻 264, 番号 35. pp. 21257-21265.
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abstract = "Factor IX B(M) Nagoya (IX Nagoya) is a natural mutant of factor IX responsible for severe hemophilia B. A patient with this mutant is characterized by a markedly prolonged ox brain prothrombin time. IX Nagoya was purified from the patient's plasma by immunoaffinity chromatography with an anti-factor IX monoclonal antibody column. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that treatment of IX Nagoya with factor XIa/Ca2+ resulted in cleavage only at the Arg145-Ala146 bond. Reversed-phase high performance liquid chromatography of a trypsin digest of IX Nagoya showed an aberrant peptide, which was further digested with proteinase Asp-N. Primary structure analysis of one of the Asp-N peptides revealed that Arg180 is replaced by Trp. An essentially complete (99{\%}) amino acid sequence of IX Nagoya was obtained by sequencing fragments derived from a lysyl endopeptidase digest in which no other substitutions in the catalytic triad or substrate binding site were found. We also found that IX Nagoya is activated by α-chymotrypsin or rat mast cell chymase by monitoring the rate of factor X activation using a fluorogenic peptide substrate in the presence of factor VII, phospholipids, and Ca2+. These results indicate that the substitution of Arg180 by Trp impairs the cleavage by factor XIa required for activation of this zymogen and that the substitution causes hemophilia B(M).",
author = "K. Suehiro and Shun-Ichiro Kawabata and T. Miyata and H. Takeya and J. Takamatsu and K. Ogata and T. Kamiya and H. Saito and Y. Niho and S. Iwanaga",
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T1 - Blood clotting factor IX B(M) Nagoya. Substitution of arginine 180 by tryptophan and its activation by α-chymotrypsin and rat mast cell chymase

AU - Suehiro, K.

AU - Kawabata, Shun-Ichiro

AU - Miyata, T.

AU - Takeya, H.

AU - Takamatsu, J.

AU - Ogata, K.

AU - Kamiya, T.

AU - Saito, H.

AU - Niho, Y.

AU - Iwanaga, S.

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N2 - Factor IX B(M) Nagoya (IX Nagoya) is a natural mutant of factor IX responsible for severe hemophilia B. A patient with this mutant is characterized by a markedly prolonged ox brain prothrombin time. IX Nagoya was purified from the patient's plasma by immunoaffinity chromatography with an anti-factor IX monoclonal antibody column. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that treatment of IX Nagoya with factor XIa/Ca2+ resulted in cleavage only at the Arg145-Ala146 bond. Reversed-phase high performance liquid chromatography of a trypsin digest of IX Nagoya showed an aberrant peptide, which was further digested with proteinase Asp-N. Primary structure analysis of one of the Asp-N peptides revealed that Arg180 is replaced by Trp. An essentially complete (99%) amino acid sequence of IX Nagoya was obtained by sequencing fragments derived from a lysyl endopeptidase digest in which no other substitutions in the catalytic triad or substrate binding site were found. We also found that IX Nagoya is activated by α-chymotrypsin or rat mast cell chymase by monitoring the rate of factor X activation using a fluorogenic peptide substrate in the presence of factor VII, phospholipids, and Ca2+. These results indicate that the substitution of Arg180 by Trp impairs the cleavage by factor XIa required for activation of this zymogen and that the substitution causes hemophilia B(M).

AB - Factor IX B(M) Nagoya (IX Nagoya) is a natural mutant of factor IX responsible for severe hemophilia B. A patient with this mutant is characterized by a markedly prolonged ox brain prothrombin time. IX Nagoya was purified from the patient's plasma by immunoaffinity chromatography with an anti-factor IX monoclonal antibody column. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that treatment of IX Nagoya with factor XIa/Ca2+ resulted in cleavage only at the Arg145-Ala146 bond. Reversed-phase high performance liquid chromatography of a trypsin digest of IX Nagoya showed an aberrant peptide, which was further digested with proteinase Asp-N. Primary structure analysis of one of the Asp-N peptides revealed that Arg180 is replaced by Trp. An essentially complete (99%) amino acid sequence of IX Nagoya was obtained by sequencing fragments derived from a lysyl endopeptidase digest in which no other substitutions in the catalytic triad or substrate binding site were found. We also found that IX Nagoya is activated by α-chymotrypsin or rat mast cell chymase by monitoring the rate of factor X activation using a fluorogenic peptide substrate in the presence of factor VII, phospholipids, and Ca2+. These results indicate that the substitution of Arg180 by Trp impairs the cleavage by factor XIa required for activation of this zymogen and that the substitution causes hemophilia B(M).

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