title: Properties and regulation of Mg2+-dependent chloroplast inorganic pyrophosphatase from Sorghum vulgare leaves.
abstract: A Mg2+ dependent inorganic pyrophosphatase from chloroplasts of Sorghum vulgare has been purified 275-fold to electrophoretic purity with an overall recovery of about 25% activity. Estimations of native and monomeric relative molecular weights by size exclusion chromatography and denaturing electrophoresis suggest that the holoenzyme is a monomer of 42 +/- 1.5 kDa. A high specificity for tetrasodium pyrophosphate (PPi) as substrate has been observed, as the other phosphoesters tested were virtually unaffected. The Mg2+:PPi ratio of 5:1 at pH 8.0 shifts to 2.5:1.0 at pH 9.0 and 10:1 at pH 7.0. None of the divalent cations tested could substitute for Mg2+. Further, in the presence of Mg2+, these divalent cations inhibit the catalytic hydrolysis of PPi. EDTA rapidly and irreversibly inactivates the purified enzyme in a biphasic manner. Of the metabolites tested, Pi and L-malate significantly inhibited the catalytic activity of the enzyme. Malate inhibits the enzyme through an allosteric mechanism. A Hill plot of this inhibition shows that at least two molecules of malate bind to each molecule of the purified enzyme. The likely physiological significance of this result is discussed.
authors.affiliation: School of Biological Sciences, Madurai Kamaraj University, India.
authors.forename: V A
authors.initials: VA
authors.lastname: Krishnan
authors.forename: A
authors.initials: A
authors.lastname: Gnanam
mesh_terms: D002413:Cations, Divalent; D002736:Chloroplasts / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008274:Magnesium / Q000502:physiology*; D008970:Molecular Weight; D014674:Plant Proteins, Dietary / Q000302:isolation & purification; D006109:Poaceae / Q000201:enzymology*; D011755:Pyrophosphatases / Q000302:isolation & purification*; D013379:Substrate Specificity
chemical_list: D002413:Cations, Divalent; D014674:Plant Proteins, Dietary; D011755:Pyrophosphatases; D008274:Magnesium