Purification and Some Properties of Inorganic Pyrophosphatase from Streptococcus faecalis1
- 1 July 1981
- journal article
- research article
- Published by Oxford University Press (OUP) in The Journal of Biochemistry
- Vol. 90 (1) , 79-85
- https://doi.org/10.1093/oxfordjournals.jbchem.a133471
Abstract
Inorganic pyrophosphatase [EC 3.6.1.1] from Streptococcus faecalis ATCC 8043 was purified to homogeneity as judged by slab gel electrophoresis in the presence and absence of sodium dodecyl sulfate. The purification consisted of the following steps: streptomycin sulfate precipitation, (NH4)2SO4 treatment in a Sepharose CL-4B column, DEAE-Sepharose CL-6B chromatography, gel filtration on Ultrogel AcA 34, and preparative slab gel electrophoresis. Gel filtration on Ultrogel AcA 34 was used to determine the molecular weight and slab gel electrophoresis in the presence of sodium dodecyl sulfate to study the subunit molecular weight. The enzyme appeared to be composed of four subunits with molecular weights of approximately 32,500. The molecular weight of the native enzyme was about 128,000. The enzyme catalyzed the hydrolysis of inorganic pyrophosphate, and no activity was found with a variety of other phosphate esters. The cation Mg2+ was required for maximum activity; Co2+ and Ca2+ supported 24% and 5.6% of the activity observed with Mg2+, respectively. 2,4,6-Trinitrobenzene sulfonic acid inhibited and reductants containing SH-groups activated the enzyme suggesting that lysine and cysteine have essential roles in the enzyme.Keywords
This publication has 20 references indexed in Scilit:
- Partial inhibition of DNA synthesis gives rise to increase in the level of inorganic pyrophosphatase in the growing cells of Escherichia coliChemico-Biological Interactions, 1977
- Fluoride inhibition of inorganic pyrophosphatase III. Dependence on the nature of substrate and metal ion cofactorBiochimica et Biophysica Acta (BBA) - Enzymology, 1977
- Fluoride inhibition of inorganic pyrophosphatase II. Isolation and characterization of a covalent intermediate between enzyme and entire substrate moleculeBiochimica et Biophysica Acta (BBA) - Enzymology, 1977
- Regulation of Intracellular Pyrophosphatase-Activity and Conservation of the Phosphoanhydride-Energy of Inorganic Pyrophosphate in Microbial MetabolismZeitschrift für Naturforschung C, 1976
- Fluoride inhibition of inorganic pyrophosphatase I. Kinetic studies in A Mg2+-PPi system using a new continuous enzyme assayBiochimica et Biophysica Acta (BBA) - Enzymology, 1976
- BIOCHEMICAL STUDIES OF BACTERIAL SPORULATION .3. INORGANIC PYROPHOSPHATASE OF VEGETATIVE CELLS AND SPORES OF BACILLUS SUBTILIS1967
- CONSTITUTIVE INORGANIC PYROPHOSPHATASE OF ESCHERICHIA COLI .I. PURIFICATION AND CATALYTIC PROPERTIES1966
- CONSTITUTIVE INORGANIC PYROPHOSPHATASE OF ESCHERICHIA COLI .2. NATURE AND BINDING OF ACTIVE SUBSTRATE AND ROLE OF MAGNESIUM1966
- A COBALT-ACTIVATED BACTERIAL PYROPHOSPHATASEJournal of Bacteriology, 1955
- PROTEIN MEASUREMENT WITH THE FOLIN PHENOL REAGENTJournal of Biological Chemistry, 1951