Phosphorolytic cleavage of diadenosine 5',5'''-P1,P4-tetraphosphate. Properties of homogeneous diadenosine 5',5'''-P1,P4-tetraphosphate alpha, beta-phosphorylase from Saccharomyces cerevisiae.
Novel enzymatic activity which splits diadenosine 5',5'''-P1,P4-tetraphosphate (Ap4A) phosphorolytically has been found in extracts from Saccharomyces cerevisiae. One of the two alpha,beta-anhydride bonds between Ap4A phosphate residues undergoes phosphorolysis, and ATP (pppA) plus ADP (ppA) are the products of the reaction according to the equation: AppppA + P*i----pppA + p*pA The reaction is dependent on the presence of divalent metal ions; Mn2+ or Mg2+ sustain the greatest rates of reaction. Among analogues of the Ap4A substrate, Ap5A and Gp4G, but not p4A and Ap3A, are substrates, and corresponding products are p4A plus ADP, and GTP plus GDP, the phosphate being incorporated into the nucleoside 5'-diphosphates. In the reactions, phosphate can be substituted with arsenate. Arsenolysis of Ap4A, Ap5A, or Gp4G leads to ATP plus AMP, p4A plus AMP, and GTP plus GMP, respectively. The name diadenosine tetraphosphate alpha,beta-phosphorylase (ADP-forming) is proposed for the new enzyme. The phosphorylase has been purified to apparent homogeneity and behaves as a single polypeptide chain of Mr = 40,000. Optimum activity of the enzyme is at pH 8.0 and the sulfhydryl groups are essential for catalysis. At saturating Ap4A, the rate constant for the reaction is 36 s-1 and the Km value for Ap4A is 60 microM (37 degrees C, 50 mM Hepes/KOH (pH 8.2), 500 microM MnCl2, 10 mM K2HPO4, 1 mM 2-mercaptoethanol, and 2% glycerol). The Km values for phosphate and arsenate are 1 and 3 mM, respectively.
Guranowski
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Blanquet
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D000227:Adenine Nucleotides / Q000378:metabolism*; D002413:Cations, Divalent / Q000494:pharmacology; D002850:Chromatography, Gel; D015226:Dinucleoside Phosphates; D004591:Electrophoresis, Polyacrylamide Gel; D009713:Nucleotidyltransferases / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D013379:Substrate Specificity
D000227:Adenine Nucleotides; D002413:Cations, Divalent; D015226:Dinucleoside Phosphates; C020733:diadenosine tetraphosphate; D009713:Nucleotidyltransferases; C044740:diadenosine 5,5'''-P(1),P(4)-tetraphosphate alpha,beta-phosphorylase