Studies of vitamin K-dependent carboxylase
Carboxylase was studied in detergent solubilized rat liver microsomes, using the peptide substrate Phe-Leu-(..gamma..-/sup 3/H)-Glu-Glu-Leu. Cleavage of the ..gamma..-C-H bond in Glu was measured as the release of /sup 3/H from this peptide to water, carboxylation was measured as the incorporation of H/sup 14/CO/sub 3/-into the peptide, and KO formation was measured by an HPLC assay. All three products could be measured simultaneously, and this system was used to examine the effects of cyanide, manganese, tetrachloropyridinol, and Boc-SerP-SerP-Leu-OMe on the separate steps of the carboxylase reaction. Vitamin K-epoxide formation was studied separately from the other reactions, and it was found that in the absence of a Glu-containing substrate, carboxylase catalyzed the uncoupled formation of KO from KH/sub 2/ and O/sub 2/. The stoichiometry of product formation (GLa, KO, and ..gamma..-protons) was measured, and the results obtained were all in agreement with the values predicted from the proposed mechanism. When all of the substrates were saturating, the stoichiometry of ..gamma..-C-H bond cleavage, carboxylation, and KO formation was 1:1:1.
- Research Organization:
- Wisconsin Univ., Madison (USA)
- OSTI ID:
- 5281553
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
59 BASIC BIOLOGICAL SCIENCES
ANIMALS
AROMATICS
BIOCHEMICAL REACTION KINETICS
BODY
CARBON 14 COMPOUNDS
CARBON-CARBON LYASES
CARBOXY-LYASES
CARBOXYLASE
CARBOXYLATION
CELL CONSTITUENTS
CHEMICAL REACTIONS
CHROMATOGRAPHY
COAGULANTS
DIGESTIVE SYSTEM
DRUGS
ENZYMES
GLANDS
HEMATOLOGIC AGENTS
HEPARIN ANTAGONISTS
ISOTOPE APPLICATIONS
KINETICS
LABELLED COMPOUNDS
LIQUID COLUMN CHROMATOGRAPHY
LIVER
LYASES
MAMMALS
MICROSOMES
ORGANIC COMPOUNDS
ORGANIC OXYGEN COMPOUNDS
ORGANOIDS
ORGANS
PEPTIDES
PROTEINS
QUINONES
RATS
REACTION KINETICS
RODENTS
SEPARATION PROCESSES
STOICHIOMETRY
SUBSTRATES
TRACER TECHNIQUES
TRITIUM COMPOUNDS
VERTEBRATES
VITAMIN K
VITAMINS