Vitamin K-dependent carboxylation. In vitro modification of synthetic peptides containing the γ-carboxylation recognition site

B. R. Hubbard, M. Jacobs, M. M.W. Ulrich, C. Walsh, B. C. Furie, B. C. Furie

Research output: Contribution to journalArticleAcademicpeer-review

Abstract

Synthetic peptides including the γ-carboxylation recognition site and acidic amino acids were compared as substrates for vitamin K-dependent γ-carboxylation by bovine liver carboxylase. The 28-residue proPT28 (proprothrombin -18 to +10) and proFIX28 (proFactor IX -18 to +10) were carboxylated with a K(m) of 3 μM. The V(max) of proPT28 was 2-3 times greater than that of proFIX28. An analog of proFIX28 that contained the prothrombin propeptide had a V(max) 2-3-fold greater than an analog of proPT28 that contained the Factor IX propeptide.proFIX28/RS-1, based upon Factor IX Cambridge, proFIX28/RQ-4, based upon Factor IX Oxford 3, and proFIX28 had equivalent K(m) and V(max) values. Analogs of proPT28 containing Ala6-Glu7 or Glu6-Ala7 were carboxylated at equivalent rates. A peptide containing Asp6-Asp7 was carboxylated at a rate of about 1% of that of Glu carboxylation. Carboxylation of peptides containing Asp6-Glu7 and Glu6-Asp7 yielded results identical with peptides containing Ala6-Glu7 and Glu6-Ala7. Carboxymethylcysteine was not carboxylated when substituted for Glu6 in a peptide containing Asp7. These results indicate that the prothrombin propeptide is more efficient in the carboxylation process than is the Factor IX propeptide, but that both propetides direct carboxylation; the γ-carboxylation recognition site does not include residues -4 and -1; aspartic acid and carboxymethylcysteine are poor substrates for the carboxylase, but aspartic acid does not inhibit the carboxylation of adjacent glutamic acids.

Original languageEnglish
Pages (from-to)14145-14150
Number of pages6
JournalJournal of Biological Chemistry
Volume264
Issue number24
Publication statusPublished - 1 Jan 1989

Cite this

Hubbard, B. R. ; Jacobs, M. ; Ulrich, M. M.W. ; Walsh, C. ; Furie, B. C. ; Furie, B. C. / Vitamin K-dependent carboxylation. In vitro modification of synthetic peptides containing the γ-carboxylation recognition site. In: Journal of Biological Chemistry. 1989 ; Vol. 264, No. 24. pp. 14145-14150.
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abstract = "Synthetic peptides including the γ-carboxylation recognition site and acidic amino acids were compared as substrates for vitamin K-dependent γ-carboxylation by bovine liver carboxylase. The 28-residue proPT28 (proprothrombin -18 to +10) and proFIX28 (proFactor IX -18 to +10) were carboxylated with a K(m) of 3 μM. The V(max) of proPT28 was 2-3 times greater than that of proFIX28. An analog of proFIX28 that contained the prothrombin propeptide had a V(max) 2-3-fold greater than an analog of proPT28 that contained the Factor IX propeptide.proFIX28/RS-1, based upon Factor IX Cambridge, proFIX28/RQ-4, based upon Factor IX Oxford 3, and proFIX28 had equivalent K(m) and V(max) values. Analogs of proPT28 containing Ala6-Glu7 or Glu6-Ala7 were carboxylated at equivalent rates. A peptide containing Asp6-Asp7 was carboxylated at a rate of about 1{\%} of that of Glu carboxylation. Carboxylation of peptides containing Asp6-Glu7 and Glu6-Asp7 yielded results identical with peptides containing Ala6-Glu7 and Glu6-Ala7. Carboxymethylcysteine was not carboxylated when substituted for Glu6 in a peptide containing Asp7. These results indicate that the prothrombin propeptide is more efficient in the carboxylation process than is the Factor IX propeptide, but that both propetides direct carboxylation; the γ-carboxylation recognition site does not include residues -4 and -1; aspartic acid and carboxymethylcysteine are poor substrates for the carboxylase, but aspartic acid does not inhibit the carboxylation of adjacent glutamic acids.",
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Vitamin K-dependent carboxylation. In vitro modification of synthetic peptides containing the γ-carboxylation recognition site. / Hubbard, B. R.; Jacobs, M.; Ulrich, M. M.W.; Walsh, C.; Furie, B. C.; Furie, B. C.

In: Journal of Biological Chemistry, Vol. 264, No. 24, 01.01.1989, p. 14145-14150.

Research output: Contribution to journalArticleAcademicpeer-review

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T1 - Vitamin K-dependent carboxylation. In vitro modification of synthetic peptides containing the γ-carboxylation recognition site

AU - Hubbard, B. R.

AU - Jacobs, M.

AU - Ulrich, M. M.W.

AU - Walsh, C.

AU - Furie, B. C.

AU - Furie, B. C.

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N2 - Synthetic peptides including the γ-carboxylation recognition site and acidic amino acids were compared as substrates for vitamin K-dependent γ-carboxylation by bovine liver carboxylase. The 28-residue proPT28 (proprothrombin -18 to +10) and proFIX28 (proFactor IX -18 to +10) were carboxylated with a K(m) of 3 μM. The V(max) of proPT28 was 2-3 times greater than that of proFIX28. An analog of proFIX28 that contained the prothrombin propeptide had a V(max) 2-3-fold greater than an analog of proPT28 that contained the Factor IX propeptide.proFIX28/RS-1, based upon Factor IX Cambridge, proFIX28/RQ-4, based upon Factor IX Oxford 3, and proFIX28 had equivalent K(m) and V(max) values. Analogs of proPT28 containing Ala6-Glu7 or Glu6-Ala7 were carboxylated at equivalent rates. A peptide containing Asp6-Asp7 was carboxylated at a rate of about 1% of that of Glu carboxylation. Carboxylation of peptides containing Asp6-Glu7 and Glu6-Asp7 yielded results identical with peptides containing Ala6-Glu7 and Glu6-Ala7. Carboxymethylcysteine was not carboxylated when substituted for Glu6 in a peptide containing Asp7. These results indicate that the prothrombin propeptide is more efficient in the carboxylation process than is the Factor IX propeptide, but that both propetides direct carboxylation; the γ-carboxylation recognition site does not include residues -4 and -1; aspartic acid and carboxymethylcysteine are poor substrates for the carboxylase, but aspartic acid does not inhibit the carboxylation of adjacent glutamic acids.

AB - Synthetic peptides including the γ-carboxylation recognition site and acidic amino acids were compared as substrates for vitamin K-dependent γ-carboxylation by bovine liver carboxylase. The 28-residue proPT28 (proprothrombin -18 to +10) and proFIX28 (proFactor IX -18 to +10) were carboxylated with a K(m) of 3 μM. The V(max) of proPT28 was 2-3 times greater than that of proFIX28. An analog of proFIX28 that contained the prothrombin propeptide had a V(max) 2-3-fold greater than an analog of proPT28 that contained the Factor IX propeptide.proFIX28/RS-1, based upon Factor IX Cambridge, proFIX28/RQ-4, based upon Factor IX Oxford 3, and proFIX28 had equivalent K(m) and V(max) values. Analogs of proPT28 containing Ala6-Glu7 or Glu6-Ala7 were carboxylated at equivalent rates. A peptide containing Asp6-Asp7 was carboxylated at a rate of about 1% of that of Glu carboxylation. Carboxylation of peptides containing Asp6-Glu7 and Glu6-Asp7 yielded results identical with peptides containing Ala6-Glu7 and Glu6-Ala7. Carboxymethylcysteine was not carboxylated when substituted for Glu6 in a peptide containing Asp7. These results indicate that the prothrombin propeptide is more efficient in the carboxylation process than is the Factor IX propeptide, but that both propetides direct carboxylation; the γ-carboxylation recognition site does not include residues -4 and -1; aspartic acid and carboxymethylcysteine are poor substrates for the carboxylase, but aspartic acid does not inhibit the carboxylation of adjacent glutamic acids.

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JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

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