Vitamin K-dependent carboxylation of peptide-bound glutamate. The active species of 'CO2' utilized by the membrane-bound preprothrombin carboxylase

J. P. Jones, E. J. Gardner, Terrance Cooper, R. E. Olson

Research output: Contribution to journalArticle

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Abstract

Vitamin K participates in the post-translational carboxylation of peptide bound glutamate to form the γ-carboxyglutamate residues of prothrombin. The reaction requires reduced vitamin K, bicarbonate, oxygen, and a membrane bound carboxylase. The active species of 'CO2,' i.e. CO2 or HCO3 -, utilized in this carboxylation was determined by the low temperature method of Filmer and Cooper (1970), taking advantage of the fact that menaquinone-2, in contrast to phylloquinone, is very active at 10°. Microsomes from livers of vitamin K-deficient rats, were incubated in the presence of cycloheximide, avidin, NADH, menaquinone-2, 1 mM acetazolamide (to inhibit carbonic anhydrase), and either 14CO2 or H14CO3 -. At 1-min intervals aliquots were removed from the reaction mixture. γ-Carboxyglutamate was isolated from these samples by ion exchange chromatography after alkaline hydrolysis. After 1 min the incorporation of 14CO2 into γ-carboxyglutamate was 8 to 10 times as great as that found with H14CO3 -. When the carbonic anhydrase inhibitor was omitted (with or without addition of exogenous carbonic anhydrase) the two incorporation curves approximated each other at a rate near that exhibited by bicarbonate alone. Similar results were obtained in a microsomal carboxylase system solubilized with Triton X-100. It is concluded that CO2 is the active species of 'CO2' initially participating in the vitamin K-dependent carboxylation of preprothrombin and that neither ATP nor biotin is required for the reaction.

Original languageEnglish (US)
Pages (from-to)7738-7742
Number of pages5
JournalJournal of Biological Chemistry
Volume252
Issue number21
StatePublished - Dec 1 1977
Externally publishedYes

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Carboxylation
Vitamin K
Vitamin K 2
Glutamic Acid
Carbonic Anhydrases
Bicarbonates
Membranes
Peptides
Vitamin K 1
Carbonic Anhydrase Inhibitors
Acetazolamide
Avidin
Ion Exchange Chromatography
Octoxynol
Prothrombin
Liver Microsomes
Cycloheximide
Biotin
Chromatography
Liver

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Vitamin K-dependent carboxylation of peptide-bound glutamate. The active species of 'CO2' utilized by the membrane-bound preprothrombin carboxylase. / Jones, J. P.; Gardner, E. J.; Cooper, Terrance; Olson, R. E.

In: Journal of Biological Chemistry, Vol. 252, No. 21, 01.12.1977, p. 7738-7742.

Research output: Contribution to journalArticle

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