Selective splitting of 3'-adenylated dinucleoside polyphosphates by specific enzymes degrading dinucleoside polyphosphates.
Several 3'-[(32)P]adenylated dinucleoside polyphosphates (Np(n)N'p*As) were synthesized by the use of poly(A) polymerase (Sillero MAG et al., 2001, Eur J Biochem.; 268: 3605-11) and three of them, ApppA[(32)P]A or ApppAp*A, AppppAp*A and GppppGp*A, were tested as potential substrates of different dinucleoside polyphosphate degrading enzymes. Human (asymmetrical) dinucleoside tetraphosphatase (EC 3.6.1.17) acted almost randomly on both AppppAp*A, yielding approximately equal amounts of pppA + pAp*A and pA + pppAp*A, and GppppGp*, yielding pppG + pGp*A and pG + pppGp*A. Narrow-leafed lupin (Lupinus angustifolius) tetraphosphatase acted preferentially on the dinucleotide unmodified end of both AppppAp*A (yielding 90% of pppA + pAp*A and 10 % of pA + pppAp*A) and GppppGp*A (yielding 89% pppG + pGp*A and 11% of pG + pppGp*A). (Symmetrical) dinucleoside tetraphosphatase (EC 3.6.1.41) from Escherichia coli hydrolyzed AppppAp*A and GppppGp*A producing equal amounts of ppA + ppAp*A and ppG + ppGp*A, respectively, and, to a lesser extent, ApppAp*A producing pA + ppAp*A. Two dinucleoside triphosphatases (EC 3.6.1.29) (the human Fhit protein and the enzyme from yellow lupin (Lupinus luteus)) and dinucleoside tetraphosphate phosphorylase (EC 2.7.7.53) from Saccharomyces cerevisiae did not degrade the three 3'-adenylated dinucleoside polyphosphates tested.
Guranowski
Andrzej
A
Departamento de Bioquímica, Instituto de Investigaciones Biomédicas Alberto Sols UAM/CSIC, Facultad de Medicina, 28029 Madrid, Spain. guranow@au.poznan.pl
Sillero
Antonio
A
Günther Sillero
María Antonia
MA
D017766:Acid Anhydride Hydrolases / Q000378:metabolism*; D002855:Chromatography, Thin Layer; D015226:Dinucleoside Phosphates / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D007700:Kinetics; D011061:Poly A / Q000378:metabolism*; D011062:Polynucleotide Adenylyltransferase / Q000378:metabolism*; D013379:Substrate Specificity
D015226:Dinucleoside Phosphates; D011061:Poly A; D011062:Polynucleotide Adenylyltransferase; D017766:Acid Anhydride Hydrolases; C020023:bis(5'-nucleosyl)tetraphosphatase (asymmetrical)