Alternate substrates and spectroscopic studies of P-enolpyruvate carboxykinase

Conference · Thu May 01 04:00:00 EDT 1986 · Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States)

OSTI ID:5151200

Ash, D E; Schramm, V L

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.

• Research Organization: Temple Univ., Philadelphia, PA
• OSTI ID: 5151200
• Report Number(s): CONF-8606151-
• Conference Information: Journal Name: Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States) Journal Volume: 45:6
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
550600 -- Medicine
59 BASIC BIOLOGICAL SCIENCES
62 RADIOLOGY AND NUCLEAR MEDICINE
ANIMALS
BIOCHEMICAL REACTION KINETICS
BODY
CARBOXYLIC ACIDS
CATIONS
CHARGED PARTICLES
CHEMICAL REACTIONS
DIGESTIVE SYSTEM
ELECTRON SPIN RESONANCE
ENZYMES
GLANDS
GLYCOLIC ACID
HYDROXY ACIDS
IONS
ISOTOPES
KINETICS
LIGHT NUCLEI
LIVER
MAGNETIC RESONANCE
MAMMALS
MONOCARBOXYLIC ACIDS
NMR SPECTRA
NUCLEI
ODD-EVEN NUCLEI
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANS
PHOSPHORUS 31
PHOSPHORUS ISOTOPES
PHOSPHORUS-GROUP TRANSFERASES
PHOSPHORYLATION
PHOSPHOTRANSFERASES
RATS
REACTION KINETICS
RESONANCE
RODENTS
SPECTRA
STABLE ISOTOPES
SUBSTRATES
TRANSFERASES
VALENCE
VERTEBRATES