Studies on the mechanism and active site of yeast inorganic pyrophosphatase
The carboxyl groups in yeast inorganic pyrophosphatase (PPase) has been studied and its role in the mechanism of pyrophosphate hydrolysis has been proved by means of kinetics, stereochemical and chemical modification studies. In addition the authors have confirmed the formation of an acylphosphate in the presence of fluoride. The presence of an intermediate in catalysis was studied by stereochemical methods and by a single turnover experiment. It was observed that ..gamma..-ATPS is a substrate in the presence of divalent metal ions. Incubation of a chiral ATPS with PPase led to formation of thiophosphate with inversion of configuration as determined by /sup 31/P NMR analysis. This indicated that only one displacement occurred at the thiophosphoryl center. Pyrophosphate hydrolysis was carried out under single turnover conditions in the presence of H/sub 2//sup 18/O. Analysis of phosphate by /sup 31/P NMR showed that oxygen uptake on PPase catalysis come directly from water as measured by the incorporation of /sup 18/O. These results provide strong evidence that PPase catalysis proceeds without intermediate formation.
- Research Organization:
- Pennsylvania Univ., Philadelphia (USA)
- OSTI ID:
- 6549961
- Resource Relation:
- Other Information: Thesis (Ph. D)
- Country of Publication:
- United States
- Language:
- English
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PHOSPHATASES
BIOCHEMICAL REACTION KINETICS
PHOSPHATES
NMR SPECTRA
PYROPHOSPHATES
METABOLISM
ATP-ASE
FLUORIDES
NUCLEAR MAGNETIC RESONANCE
OXYGEN 18
PHOSPHORUS 31
STEREOCHEMISTRY
YEASTS
ACID ANHYDRASES
ENZYMES
ESTERASES
EVEN-EVEN NUCLEI
FLUORINE COMPOUNDS
FUNGI
HALIDES
HALOGEN COMPOUNDS
HYDROLASES
ISOTOPES
KINETICS
LIGHT NUCLEI
MAGNETIC RESONANCE
MICROORGANISMS
NUCLEI
ODD-EVEN NUCLEI
OXYGEN COMPOUNDS
OXYGEN ISOTOPES
PHOSPHOHYDROLASES
PHOSPHORUS COMPOUNDS
PHOSPHORUS ISOTOPES
PLANTS
REACTION KINETICS
RESONANCE
SPECTRA
STABLE ISOTOPES
550500* - Metabolism
550200 - Biochemistry