Riboside triphosphates

Chromatin isolated by this method, as the initial experiment showed, was capable of synthesizing RNA from riboside triphosphates (Huang el at., 1960 Bonner et al., 1961), and the presence of all four riboside triphosphates (adenyl, guanyl, cytosyl, and uridyl) was necessary for this synthesis. This synthesis was DNA-dependent. 

Figure 14 IPT and tRNA-IPT reaction for primary cytokinin biosynthesis. Circles and A in tRNA show anticodon and adenosine, respectively. iPRMP, iP riboside 5 -monophosphate iPRDP, iP riboside 5 -diphosphate iPRTP, iP riboside 5 -triphosphate tZRMP, tZ riboside 5 -monophosphate PP, diphosphoric acid.

Figure 15 Current model of isoprenoid cytokinin biosynthesis pathway in higher plants. tZRDP, tZ riboside 5 -diphosphate tZRTP, tZ riboside 5 -triphosphate DZRMP, DZ riboside 5 -monophosphate cZRMP, cZ riboside 5 -monophosphate ...

Fig. 11.1.4. Separation of uracil and 5-fluorouracil bases, nucleosides and nucleotides by reversed phase ion-pair HPLC. Chromatographic conditions column, Bondapak Cig (300 x 4 mm) mobile phase, (from 0-30 min) 0.1 mM tetrabutylammonium hydrogen sulphate (Cjg), 2.5 mM tetraethylammonium bromide (Cg) and 2% methanol in 2 mM sodium acetate, 1.5 mM phosphate buffer, pH 6.0 (Buffer A) (from 30-50 min) Buffer A-i-30 mM phosphate detection, UV at 254 nm. Peaks FU, fluorouracil FUR, fluorouracU riboside/ FUdR, fluorouracil deoxyriboside FUMP, fluorouridine 5 -monophosphate 5 dFUR, 5 -deoxyfluorouracil riboside FdUMP, deoxyfluorouri-dine monophosphate UDPG, uridine diphosphoglucose UDP, uridine diphosphate dUDP, deoxyuridine monophosphate UTP, uridine triphosphate. Reproduced from Au et al. (1982), with permission.

In spite of their biological and agricultural importance, only in the last decade the basic molecular mechanism of their biosynthesis and signal transduction have been elucidated. The first step in the isoprenoid CK biosynthesis is the M-prenylation of adenosine-5 -phosphate (AMP, ADP, or ATP) with DMAPP or hydroxymethylbutenyl diphosphate (HMBDP) catalyzed by adenosine phosphate-isopentenyltransferase (IPT). Plant IPTs preferentially utilize ATP or ADP as the isoprenoid acceptors to form isopentenyl riboside 5 -triphosphate (iPRTP) and... 

The terms nucleoside and nucleotide in the strictest sense refer to N- lycosides and phosphorylated N-glycosides, respectively, derived from nucleic acids. The term is now used, however, in several broader ways. Thus, adenosine triphosphate (ATP) is not derived from nucleic acids, but is quite legitimately a nucleotide through its relation to adenosine monophosphate (AMP), which is so derived. Other N-ribosides, such as nicotinamide mononucleotide (NMN), are called nucleotides only by extension and anal-... 

From the 5 -monophosphates the corresponding 5 -diphosphates and 5 -tri-phosphates, e.g., adenosine triphosphate (ATP) and guanosine triphosphate (GTP), the unphosphorylated ribosides inosine, adenosine, and guanosine, as well as the free purines hypoxanthine, adenine, and guanine may be formed. 

Morphologists postulated for many years that the transfer of information from DNA to protein involves a form of nuclear RNA that migrates from nucleus to cytoplasm. This hypothesis found some concrete support as molecular biology developed. At first, it was found that a small fraction of bacterial RNA has a base composition identical to DNA in bacteria infected with T phage. The synthesis of DNA-depend-ent RNA was demonstrated in bacterial preparations and in mammalian nuclei. Triphosphate ribosides (UTP, GTP, CTP, and ATP) are required precursors for the synthesis of the new polynucleotide. Double-stranded DNA serves as a template in the reaction, and an RNA polymerase catalyzing the polymerization of the ribonucleotide was partially purified from both the bacterial and mammalian systems. 

Bios5mthetic pathways of naturally occurring cytokinins are illustrated in Fig. 29.5. The first step of cytokinin biosynthesis is the formation of A -(A -isopentenyl) adenine nucleotides catalyzed by adenylate isopentenyltransferase (EC 2.5.1.27). In higher plants, A -(A -isopentenyl)adenine riboside 5 -triphosphate or A -(A -isopentenyl)adenine riboside 5 -diphosphate are formed preferentially. In Arabidopsis, A -(A -isopentenyl)adenine nucleotides are converted into fraws-zeatin nucleotides by cytochrome P450 monooxygenases. Bioactive cytokinins are base forms. Cytokinin nucleotides are converted to nucleobases by 5 -nucleotidase and nucleosidase as shown in the conventional purine nucleotide catabolism pathway. However, a novel enzyme, cytokinin nucleoside 5 -monophosphate phosphoribo-hydrolase, named LOG, has recently been identified. Therefore, it is likely that at least two pathways convert inactive nucleotide forms of cytokinin to the active freebase forms that occur in plants [27, 42]. The reverse reactions, the conversion of the active to inactive structures, seem to be catalyzed by adenine phosphoiibosyl-transferase [43] and/or adenosine kinase [44]. In addition, biosynthesis of c/s-zeatin from tRNAs in plants has been demonstrated using Arabidopsis mutants with defective tRNA isopentenyltransferases [45]. 

Fig. 29.5 The two cytokiiim biosynthesis and activation pathways. DMAPP dimethylallyl diphosphate, iP A/ -(A -isopenteaiyl)adenme, iPRMP iP riboside S -mraiophosphate, tZ trans-zeatin, tZRTP tZ riboside 5 -triphosphate, tZRDP tZ riboside 5 -diphosphate, tZRMP tZ riboside 5 -monophosphate, cZ cis-zeatin, cZRMP cZ riboside 5 -monophosphate, cZR cZ riboside. Enzymes iPT isopentenyltransfoase, CYP735A cytochrome P450 CYP735A, tRNA-iPT transfer RNA isopentenyltransferase, LOG lonely guy, cytokinin nucleoside 5 -monophosphate phosphor-ibohydrolase (After Hirose et al. [26])...

Ecto-nucleotide polyphosphatase. This enzyme was first observed in nucleated erythrocytes (WENKSTERN ENGELHARDT, 1957) generally, it is described as a Ca +-ATPase or Mg -ATPase (adenosinetriphosphate-5 -phosphoester hydrolase, EC 3.6.1.3). The enzyme has a high specificity for they -phosphate of nucleoside-triphosphates but a broad specificity for the riboside moiety. It is not quite clear if extracellular ADP is hydrolyzed by this enzyme, but the current evidence favors the existence of a separate ecto-ADPase. 

Bochner B R, Ames B N 1982 ZTP (5-amino 4-imidazole carboxamide riboside 5 -triphosphate) A proposed alarmone for 10-formyltetrahydrofolate deficiency. Cell 29 929-937... 

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