Enzymatic synthesis of ribonucleoside-5′-phosphates from some N6-substituted adenosines

Dorée, Marcel; Terrine, Claude

doi:10.1016/0031-9422(73)85008-3

Cited by 20 publications

(7 citation statements)

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“…All attempts to find Ado phosphorylase and CK riboside phosphorylase activities in plants suggest that they are normally present at undetectably low levels [45,46]. Negative results are much less likely to be reported, therefore it is noteworthy that, from a careful study of the close potato relative tomato, Ado phosphorylase was the only known purine interconverting enzyme activity that Burch and Stuchbury [45] could not detect.…”

Section: Stckp1 Shows a Preference For Ck Over Ade Substratesmentioning

confidence: 81%

A purine nucleoside phosphorylase in Solanum tuberosum L. (potato) with specificity for cytokinins contributes to the duration of tuber endodormancy

Bromley

Warnes

Newell

et al. 2014

Biochemical Journal

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Section: Stckp1 Shows a Preference For Ck Over Ade Substratesmentioning

confidence: 81%

A purine nucleoside phosphorylase in Solanum tuberosum L. (potato) with specificity for cytokinins contributes to the duration of tuber endodormancy

Bromley

Warnes

Newell

et al. 2014

Biochemical Journal

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“…Both F-I and F-Il 5'-nucleotidases had a pH optimum around 7 with either AMP or i6Ado-5'-P as a substrate; these data were obtained with 100 mm Tris-maleate buffer at pH values from 5 to 7, and with 100 mm Tris-HCl buffer at pH 7 to 9. The results are illustrated in Figure 3 Dephosphorylation Time Course.…”

Section: Rechromatography Of the F-i And F-ii 5'-nucleotidases Onmentioning

confidence: 84%

“…The cytokinin ribonucleotide can be formed from cytokinin base (3,11,18,20), the ribonucleoside (3,7,17,18) or turnover of cytokinin-containing tRNA (13,22). Alternatively, the ribonucleotide i6Ado-5'-P2 can be synthesized by a de novo pathway from the simple metabolites AMP and A2-isopentenylpyrophosphate (6, 24).…”

Section: Introductionmentioning

confidence: 99%

“…Phenylphosphate and 32-AMP are not substrates for the enzymes. It is proposed that dephosphorylation of cytokinin nucleotide by cytosol 5'-nucleotidases may play an important role in regulating levels of "active cytokinin" in plant cells.The occurrence of cytokinin nrbonucleotide and cytokinin ribonucleoside in plant cells is well documented (2,4,5,10,11).The cytokinin ribonucleotide can be formed from cytokinin base (3,11,18,20), the ribonucleoside (3,7,17,18) or turnover of cytokinin-containing tRNA (13,22). Alternatively, the ribonucleotide i6Ado-5'-P2 can be synthesized by a de novo pathway from the simple metabolites AMP and A2-isopentenylpyrophosphate (6, 24).…”

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confidence: 99%

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Metabolism of Cytokinin

Chen

Kristopeit²

1981

Plant Physiol.

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Two forms (F-I and F-II) of 5'-nucleotidases (5'-ribonucleotide phosphohydrolase, EC 3.1.3.5) which catalyze the dephosphorylatlon of N-(A2-lsopentenyl)adenosine S'-monophosphate and AMP to form the corresponding nucleosides were partially purified from the cytosol of wheat (Ticwum aesiwm) germ. Both the F-I (molecular weight, 57,000) and F-II (molecular weight, 110,000) 5'-nucleotidases dephosphorylate the ribonucleotides at an optimum pH of7. The Km values for the cytokinin nucleotide are 3.5 micromolar (F-I enzyme) and 12.8 micromolar (F-II enzyme) in 100 millmolar Tris-maleate buffer (pH 7) at 37 C. The F-I enzyme is less rapidly inactivated by heating than is the F-II enzyme. Both nucleotidases hydrolyze purine ribonucleoside S'-phosphates, AMP being the preferred substrate. N6-(A2-isopentenyl)Adenosine 5'-monophosphate is hydrolyzed at a rate 72 and 86% that of AMP by the F-I and F-II nucleotides, respectively. Phenylphosphate and 32-AMP are not substrates for the enzymes. It is proposed that dephosphorylation of cytokinin nucleotide by cytosol 5'-nucleotidases may play an important role in regulating levels of "active cytokinin" in plant cells.The occurrence of cytokinin nrbonucleotide and cytokinin ribonucleoside in plant cells is well documented (2,4,5,10,11).The cytokinin ribonucleotide can be formed from cytokinin base (3,11,18,20), the ribonucleoside (3,7,17,18) or turnover of cytokinin-containing tRNA (13,22). Alternatively, the ribonucleotide i6Ado-5'-P2 can be synthesized by a de novo pathway from the simple metabolites AMP and A2-isopentenylpyrophosphate (6, 24). In the de novo biosynthetic pathway using a crude enzyme preparation, cytokinin ribonucleoside was also formed. Thus, 5'-nucleotidases which are capable of dephosphorylating the ribonucleotide may be contained in the crude enzyme preparation. The 5'-nucleotidase has been isolated from animal cell membrane (8, 9, 14) and cytosol3 (19). Although the precise physiological function of this enzyme system in plant and animal cells is still unclear, the 5'-nucleotidase may play a role in the regulation of cytokinin metabolism.We describe here the partial purification of 5'-nucleotidase from wheat germ cytosol, the properties of the enzyme, and kinetics of 1 This work was supported by National Science Foundation Research Grant PCM 79 03832 (to C.-M. C).2Abbreviations: i6Ado-5'-P, N6-(A2-isopentenyl)adenosine 5'-monophosphate; Ade, adenine; Ado, adenosine; i6Ado, N6-(A2-isopentenyl)adenosine.3 The term "cytosol" is used only in distinction to "membrane bound." The extraction procedures do not rigorously preclude the possibility of contamination by nucleus enzymes. the dephosphorylation of cytokinin ribonucleotide by this enzyme system. MATERIALS AND METHODS CHEMICALS AND ENZYMESAdo, AMP, i6Ado, concanavalin A (grade IV), carbowax (mol wt, 6,000), 5'-nucleotidase (Crotalus adamteus venom), and wheat (Triticum aestivum) germ were from Sigma; i6Ado-5'-P was from P-L Biochemical Co., and [8-'4C]Ado (59 mCi/mmol) was from Amersham-Searle Corp....

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“…The metabolism of cytokinins has been studied in a variety of plant tissues (2-4, 7-9, 11, 13, 15) and some of the enzymes which regulate cytokinin metabolism have also been isolated (1,6). One of the major metabolites formed from the cytokinin base is the corresponding ribonucleoside.…”

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confidence: 99%

Metabolism of Cytokinin

Chen¹,

Petschow²

1978

Plant Physiol.

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As part of the study of cytokinin metabolic pathways, an enzyme, adenosine phosphorylase (EC 2.4.2.-), which catalyzed the ribosylation of N6(A2-isopentenyl)adenine, N6-fUrfUryladenine, and adenine to form the corresponding nucleosides, was partially purifi from wheat (Triticum aestvum) germ. The pH optimum for the ribosylation of the cytokinins and adenine was from 6.5 to 7.8; for guanine and hypoxanthine it was from 7.0 to 8.5. At pH 7.2 (63 mfllimolar N-2-hydroxyethyl piperazine-N'-ethanesulfonic acid) and 37 C the Km for N -(A2-isopentenyl)adenine was 57.1 micromolar, NO-furfuryladenine, 46.5 micromolar, adenine, 32.2 micromolar, and the V,,= for N6-(A2-isopentenyl)adenWne, N-fUrfUryladenine, and adenine were 134.7, 137.1, and 193.1 nanomoles per millim protein per minute, respectively. The equilibrium constants of the phosphorolysis of N-(A2-isopentenyl)adenosine and adenosine by this enzyme indicated that the reaction strongly favored nucleoside formation. This enzyme was shown to be distinct from inosine-guanosine phosphorylase based on the differences in the Sephadex G-100 gel ffitration behaviors, pH optima, and the product and p-hydroxymercuribenzoate inhibitor studies. These results suggest that adenosine phosphorylase may play a significant role in the regulation of cytokinin metabolism.The metabolism of cytokinins has been studied in a variety of plant tissues (2-4, 7-9, 11, 13, 15) and some of the enzymes which regulate cytokinin metabolism have also been isolated (1, 6). One of the major metabolites formed from the cytokinin base is the corresponding ribonucleoside. Purine nucleoside phosphorylase (purine nucleoside, orthophosphate ribosyl transferase, EC 2.4.2.1) which catalyzes a reversible phosphorolysis of purine nucleosides is generally believed to be inactive toward Ado2 (14,16,19) and cytokinin nucleoside. One of the possible pathways by which the cytokinin base can be converted to the corresponding ribonucleoside is the phosphoribosylation of the cytokinin base to the ribonucleotide and then conversion to the corresponding ribonucleoside by 5'-nucleotidase (1). Senesi et al. (17) recently reported that in Bacillus subtilis adenosine phosphorylase (EC 2.4.2.-) is distinct from purine nucleoside phosphorylase and that this enzyme catalyzes the formation of Ado from Ade. Adenosine phosphorylase activity was also found in some species of mycoplasmas (10) and mammalian cells (5).This paper presents the first direct evidence that in plant cells the ribosylation of cytokinin base and Ade is catalyzed by adenosine phosphorylase, and that this enzyme is distinct from inosineguanosine phosphorylase. The equilibrium constant of the ribo- Chromatograms were cut into 1-cm sections and placed in vials containing scintillation fluid (4). Radioactivity was measured in a Nuclear-Chicago Unilux II scintillation system. Counting efficiency of paper chromatogram sections was 74% for 14C. A Cary model 14 spectrophotometer was used to measure the quantity of purine base and its corresponding riboside. P...

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Enzymatic synthesis of ribonucleoside-5′-phosphates from some N6-substituted adenosines

Cited by 20 publications

References 5 publications

A purine nucleoside phosphorylase in Solanum tuberosum L. (potato) with specificity for cytokinins contributes to the duration of tuber endodormancy

A purine nucleoside phosphorylase in Solanum tuberosum L. (potato) with specificity for cytokinins contributes to the duration of tuber endodormancy

Metabolism of Cytokinin

Metabolism of Cytokinin

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