Diadenosine Triphosphate Research Articles

The presence of diadenosine 5′,5‴-P1,P3-triphosphate (Ap3A) in human platelets

Diadenosine triphosphate (Ap3A) has been identified and quantified in human platelets using a coupled enzymatic assay specific for Ap3A, after fractionation of acidic extracts with high-performence liquid chromatography. Upon thrombin-induced aggregation, Ap3A is released together with the homologue diadenosine tetraphosphate (Ap4A).Extracts of human platelets do also contain enzymatic activities that degrade diadenosine tetraphosphate as well as diadenosine triphosphate. These enzymes, however, are not released during thrombin-induced aggregation of the platelets.

Diadenosine tetraphosphatase from human leukemia cells. Purification to homogeneity and partial characterization.

Diadenosine tetraphosphatase, an enzyme splitting diadenosine tetraphosphate to AMP and ATP, has been purified to apparent homogeneity from a permanent cell line derived from a leukemic child. The purification procedure consisted of fractionation by ammonium sulfate precipitation, followed by Sephacryl 200 and DEAE-cellulose chromatography, and finally a differential membrane filtration. The enzyme is a single polypeptide chain of Mr = 17,500 as determined by gel electrophoresis in the presence of sodium dodecyl sulfate. The apparent molecular weight of the native enzyme was calculated as 20,000 from gel filtration data. The apparent Km for Ap4A was 0.5 microM as determined by two independent kinetic assays. None of the following compounds were substrates of the enzyme: diadenosine triphosphate, NAD, nucleoside 5'-phosphates (AMP, ATP, GDP, GTP, and UTP). The enzyme had optimal activity in the presence of 1 mM Mg2+, showing no activity in the presence of EDTA.

Diadenosine triphosphate splitting by rat liver extracts

A splitting activity on diadenosine triphosphate has been found in rat liver. One of the products of the cleavage is ADP. A Km of 10 μM has been found. This activity on diadenosine triphosphate seems to be specific as diadenosine tetraphosphate, a nucleotide previously described by others to occur in rat liver at very low concentration, is not a substrate of the reaction. The occurrence of diadenosine triphosphate in rat liver has not been so far reported, but a dinucleoside triphosphate structure has been described at the 5′ end of certain mRNAs. The possibility that this enzymatic activity may be involved in the hydrolysis of diadenosine triphosphate or in the processing of mRNAs is suggested.

Diguanosinetetraphosphatase from Rat Liver: Activity on Diadenosine Tetraphosphate and Inhibition by Adenosine Tetraphosphate

The hydrolysis of diadenosine tetraphosphate, a compound previously described by others to occur in liver at concentrations of around 0.1 mu M, is carried out by a specific enzyme. This enzyme has been partially purified from rat liver extracts, and the following properties have been found. The Km value for diadenosine tetraphosphate is 2 mu M; the products of hydrolysis are ATP and AMP; the Km value for diguanosine tetraphosphate is 2 mu M; none of the following substances were substrates of the enzyme: diadenosine triphosphate, diguanosine di and triphosphates, adenosine tetraphosphate, ATP, ADP, NAD+, NADP+ and bis-p-nitrophenylphosphate. Cyclic AMP was not an inhibitor of the reaction. The enzyme requires Mg2+ ions, is maximally active at a pH value of approximately 8, and has a molecular weight of 22000 as estimated by filtration on Sephadex G-100. The activation energy of the reaction was of 10250 cal times mol-1 (42886 J times mol-1). Particularly striking is the inhibition by adenosine tetraphosphate (Ki equals 48 nM) and guanosine tetraphosphate (Ki equals 14 nM). Other nucleotides tested were also competitive inhibitors with Ki values in the 10--100 mu M range.

Enzymatic synthesis of diadenosine tetraphosphate and diadenosine triphosphate with a purified lysyl-sRNA synthetase

Enzymatic synthesis of diadenosine tetraphosphate and diadenosine triphosphate with a purified lysyl-sRNA synthetase