skip to main content
OSTI.GOV title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: Alternate substrates and spectroscopic studies of P-enolpyruvate carboxykinase

Abstract

P-enolpyruvate carboxykinase (PEPCK) from rat liver cytosol has been shown to catalyze the GTP-dependent phosphorylation of several ..cap alpha..-hydroxy carboxylic acids. Glycolate, L-lactate, and L-glycerate are phosphorylated on the ..cap alpha..-oxygen by PEPCK. The products of the reactions have been characterized by their /sup 31/P NMR spectra. Glycolate gives a higher maximal velocity than does either L-lactate or L-glycerate, and with sufficient periods of incubation GTP is quantitatively converted to GDP and the corresponding phosphorylated products in each reaction. The reactions required a divalent cation as an obligatory cofactor and the velocities of the reactions increase between pH 7 and 8. In contrast to the analogous reactions catalyzed by pyruvate kinase, PEPCK does not catalyze the phosphorylation of ..beta..-hydroxypyruvate or the bicarbonate-dependent phosphorylations of fluoride ion or hydroxylamine. Purified PEPCK normally requires ..mu..molar concentrations of Mn(II) for full activity in the presence of mmolar Mg(II). The paramagnetic vanadyl (VO/sup 2 +/) ion has been shown to bind to PEPCK as a replacement for Mn(II). Saturation of the enzyme occurs near a ratio of 1.0 VO/sup 2 +//enzyme catalytic site. Although VO/sup 2 +/ does not support catalysis, EPR experiments have shown that ternary complexes of PEPCK-VO/sup 2 +/ with MgGTPmore » or the potent inhibitor oxalate are readily formed. These results show the utility of VO/sup 2 +/ as a paramagnetic probe in studies of the metal ion binding site of PEPCK.« less

Authors:
;
Publication Date:
Research Org.:
Temple Univ., Philadelphia, PA
OSTI Identifier:
5151200
Report Number(s):
CONF-8606151-
Journal ID: CODEN: FEPRA
Resource Type:
Conference
Journal Name:
Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States)
Additional Journal Information:
Journal Volume: 45:6; Conference: 76. annual meeting of the Federation of American Society for Experimental Biology, Washington, DC, USA, 8 Jun 1986
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; 62 RADIOLOGY AND NUCLEAR MEDICINE; PHOSPHOTRANSFERASES; BIOCHEMICAL REACTION KINETICS; CATIONS; ELECTRON SPIN RESONANCE; GLYCOLIC ACID; LIVER; NMR SPECTRA; PHOSPHORUS 31; PHOSPHORYLATION; RATS; SUBSTRATES; VALENCE; ANIMALS; BODY; CARBOXYLIC ACIDS; CHARGED PARTICLES; CHEMICAL REACTIONS; DIGESTIVE SYSTEM; ENZYMES; GLANDS; HYDROXY ACIDS; IONS; ISOTOPES; KINETICS; LIGHT NUCLEI; MAGNETIC RESONANCE; MAMMALS; MONOCARBOXYLIC ACIDS; NUCLEI; ODD-EVEN NUCLEI; ORGANIC ACIDS; ORGANIC COMPOUNDS; ORGANS; PHOSPHORUS ISOTOPES; PHOSPHORUS-GROUP TRANSFERASES; REACTION KINETICS; RESONANCE; RODENTS; SPECTRA; STABLE ISOTOPES; TRANSFERASES; VERTEBRATES; 550201* - Biochemistry- Tracer Techniques; 550600 - Medicine

Citation Formats

Ash, D E, and Schramm, V L. Alternate substrates and spectroscopic studies of P-enolpyruvate carboxykinase. United States: N. p., 1986. Web.
Ash, D E, & Schramm, V L. Alternate substrates and spectroscopic studies of P-enolpyruvate carboxykinase. United States.
Ash, D E, and Schramm, V L. Thu . "Alternate substrates and spectroscopic studies of P-enolpyruvate carboxykinase". United States.
@article{osti_5151200,
title = {Alternate substrates and spectroscopic studies of P-enolpyruvate carboxykinase},
author = {Ash, D E and Schramm, V L},
abstractNote = {P-enolpyruvate carboxykinase (PEPCK) from rat liver cytosol has been shown to catalyze the GTP-dependent phosphorylation of several ..cap alpha..-hydroxy carboxylic acids. Glycolate, L-lactate, and L-glycerate are phosphorylated on the ..cap alpha..-oxygen by PEPCK. The products of the reactions have been characterized by their /sup 31/P NMR spectra. Glycolate gives a higher maximal velocity than does either L-lactate or L-glycerate, and with sufficient periods of incubation GTP is quantitatively converted to GDP and the corresponding phosphorylated products in each reaction. The reactions required a divalent cation as an obligatory cofactor and the velocities of the reactions increase between pH 7 and 8. In contrast to the analogous reactions catalyzed by pyruvate kinase, PEPCK does not catalyze the phosphorylation of ..beta..-hydroxypyruvate or the bicarbonate-dependent phosphorylations of fluoride ion or hydroxylamine. Purified PEPCK normally requires ..mu..molar concentrations of Mn(II) for full activity in the presence of mmolar Mg(II). The paramagnetic vanadyl (VO/sup 2 +/) ion has been shown to bind to PEPCK as a replacement for Mn(II). Saturation of the enzyme occurs near a ratio of 1.0 VO/sup 2 +//enzyme catalytic site. Although VO/sup 2 +/ does not support catalysis, EPR experiments have shown that ternary complexes of PEPCK-VO/sup 2 +/ with MgGTP or the potent inhibitor oxalate are readily formed. These results show the utility of VO/sup 2 +/ as a paramagnetic probe in studies of the metal ion binding site of PEPCK.},
doi = {},
url = {https://www.osti.gov/biblio/5151200}, journal = {Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States)},
number = ,
volume = 45:6,
place = {United States},
year = {1986},
month = {5}
}

Conference:
Other availability
Please see Document Availability for additional information on obtaining the full-text document. Library patrons may search WorldCat to identify libraries that hold this conference proceeding.

Save / Share: