title: Dinucleoside polyphosphates stimulate the primer independent synthesis of poly(A) catalyzed by yeast poly(A) polymerase.
abstract: Novel properties of the primer independent synthesis of poly(A), catalyzed by the yeast poly(A) polymerase are presented. The commercial enzyme from yeast, in contrast to the enzyme from Escherichia coli, is unable to adenylate the 3'-OH end of nucleosides, nucleotides or dinucleoside polyphosphates (NpnN). In the presence of 0.05 mm ATP, dinucleotides (at 0.01 mm) activated the enzyme velocity in the following decreasing order: Gp4G, 100; Gp3G, 82; Ap6A, 61; Gp2G, 52; Ap4A, 51; Ap2A, 41; Gp5G, 36; Ap5A, 27; Ap3A, 20, where 100 represents a 10-fold activation in relation to a control without effector. The velocity of the enzyme towards its substrate ATP displayed sigmoidal kinetics with a Hill coefficient (nH) of 1.6 and a Km(S0.5) value of 0.308 +/- 0.120 mm. Dinucleoside polyphosphates did not affect the maximum velocity (Vmax) of the reaction, but did alter its nH and Km(S0.5) values. In the presence of 0.01 mm Gp4G or Ap4A the nH and Km(S0.5) values were (1.0 and 0.063 +/- 0.012 mm) and (0.8 and 0.170 +/- 0.025 mm), respectively. With these kinetic properties, a dinucleoside polyphosphate concentration as low as 1 micro m may have a noticeable activating effect on the synthesis of poly(A) by the enzyme. These findings together with previous publications from this laboratory point to a potential relationship between dinucleoside polyphosphates and enzymes catalyzing the synthesis and/or modification of DNA or RNA.
authors.affiliation: Departamento de Bioquímica, Instituto de Investigaciones Biomédicas Alberto Sols UAM/CSIC, Facultad de Medicina, Madrid, Spain.
authors.forename: María A Günther
authors.initials: MA
authors.lastname: Sillero
authors.forename: Anabel
authors.initials: A
authors.lastname: De Diego
authors.forename: Hugo
authors.initials: H
authors.lastname: Osorio
authors.forename: Antonio
authors.initials: A
authors.lastname: Sillero
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D002384:Catalysis / Q000187:drug effects; D002474:Cell-Free System; D015226:Dinucleoside Phosphates / Q000737:chemistry* / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D004926:Escherichia coli; D007700:Kinetics; D011061:Poly A / Q000096:biosynthesis / Q000737:chemistry*; D011062:Polynucleotide Adenylyltransferase / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism; D012441:Saccharomyces cerevisiae; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D015226:Dinucleoside Phosphates; D029701:Saccharomyces cerevisiae Proteins; D011061:Poly A; D000255:Adenosine Triphosphate; D011062:Polynucleotide Adenylyltransferase