Nucleosides phosphorolysis

As already stated, cells would appear to have the means of converting bases into deoxyribonucleotides through the reversibility of nucleoside phosphorolysis, providing deoxyribose 1-phosphate was available. It will be seen below that the coupled activities of deoxyriboaldolase and phos-phoribomutase (enzymes involved in degradation of deoxyribose phosphates) will generate deoxyribose 1-phosphate in vitro under certain conditions and, therefore, these enzymes represent a possible cellular source of this compound. However, it has not been demonstrated that any contribution to the formation of deoxyribonudeoside phosphates is made by... 

Previously constructed recombinant strain E. coli BM-D6 produced homologous purine nucleoside phosphorylase (PurNPase) acting (like PyrNPase) as a key catalyst transforming pyrimidine nucleosides into modified purine nucleosides via enz5miatic transglycosylation reaction [7]. PurNPase catalyzes stereoselective reaction of intermediate a-D-pentofuranose-1-phosphate (product of pyrimidine nucleoside phosphorolysis mediated by PyrNPase) condensation with purine heterocyclic base leading to formation of modified purine nucleoside. 

PNP catalyzes the phosphorolysis of purine nucleosides and deoxynucleosides in mammalian cells. The absence of PNP interferes with the proper degradation of 2 -deoxyguanosine, leading to an imbalance of cellular levels of deoxynucleotides... 

Phosphorolysis of the glycosyl esters of sugar nucleotides leads to the corresponding nucleoside 5 -pyrophosphate and glycosyl phosphate. A wheat-germ enzyme specific for adenosine 5 -(a-D-glucopy-ranosyl pyrophosphate),462 and a yeast enzyme specific for guanosine 5 -(a-D-mannopyranosyl pyrophosphate),463,464 are known. 

Arsenate similarly replaces phosphate in various phosphorolysis reactions, so that sucrose phosphorylase catalyzes the hydrolysis of sucrose in its presence (23), potato phosphorylase can hydrolyze amylose and amylopectin (24), nucleoside phosphorylase can hydrolyze inosine... 

Phosphorolysis of ribonucleic acid with polynucleotide phosphorylase gives a mixture of the diphosphates of the four common nucleosides, which are transformed into triphosphates with enolpyruvate phosphate and pyruvate kinase. This mixture may be used as such as a source of uridine triphosphate in the preparation of the nucleotide-sugar uridine 5 -(a-D-glucopy-ranosyl diphosphate) ( uridine-diphosphate-glucose, UDP-Glc), or as a... 

IMP is cleaved by phosphorolysis of the nucleoside to yield hypoxanthine and ribose-1-phosphate. (This reaction is similar to the phosphorolysis of glycogen by glycogen phosphorylase.)... 

A close look at this reaction reveals that in the opposite direction, the reaction is of the phosphorolysis type. For this reason, the enzymes catalyzing the reaction with ribose-l-phosphate are called phosphorylases, and they also participate in nucleic acid degradation pathways. Purine nucleoside phosphorylases thus convert hypoxanthine and guanine to either inosine and guanosine if ribose-l-phosphate is the substrate or to deoxyinosine and deoxyguanosine if deoxyribose-1-phosphate is the substrate. Uridine phosphorylase converts uracil to uridine in the presence of ribose-l-phosphate, and thymidine is formed from thymine and deoxyribose-l-phosphate through the action of thymidine phosphorylase. 

Figure 7-7. Purine nucleoside phosphorylase (PNP) catalyzed phosphorolysis is shown...

Nucleoside phosphorylases catalyze the reversible phosphorolysis in nucleosides and the transferase reaction involving purine or pyrimidine bases [42]. Scheme... 

Purine nucleoside phosphorylase (PNP), which catalyzes the reversible phosphorolysis of ribonucleosides to ribose 1 -phosphate and the free base (Scheme 7), plays an important role in purine nucleoside metabolism and in T-ceU development (59, 60). PNP inhibitors have therapeutic utility for selective destruction of T cells in T-ceU leukemias and T-cell lymphomas as well as in the treatment of T-cell-mediated autoimmune diseases and for the suppression of the post-organ transplant T-ceU response (61). 

Purine nucleosides are cleaved by the action of purine nucleoside phosphorylase with the liberation of ribose 1-phosphate (Kl, PI). The enzyme is apparently specific for purines. The material from erythrocytes catalyzes the phosphorolysis of purine but not pyrimidine nucleosides (T6.) Purine phosphorylase activity is found widespread in nature and in many animal tissues (FIO). Friedkin and Kalckar investigated an enzyme capable of cleaving purine deoxynucleosides to the aglycone and deoxy-ribose 1-phosphate. They concluded that the enzyme was identical to that which splits purine ribonucleosides (F8, F9). This enzyme is capable of degrading inosine, xanthosine, and guanosine to forms readily attacked by other enzymes. In so doing, it permits living cells to retain the ribose and deoxyribose moieties. 

Nucleotide C-N bond hydrolysis and phosphorolysis at the monomer level form part of purine salvage pathways and their mechanisms have been intensively investigated for pharmacological reasons. Humans can biosynthesise purine nucleosides de novo, whereas protozoal parasites such as those causing bilharzia and Chagas disease rely on hydrolysis of preformed nucleosides from the host. Finally, a series of ribosyl transfers from NAD are important in the modification of proteins by pathogens and have been studied extensively. 

Methods for the s)mthesis of nucleotides may be divided into four categories (1) phosphorylation of available hydroxyl groups of selectively protected nucleosides with phosphorylating agents, (2) phosphorolysis of activated nucleosides, and (3) condensation of a suitably protected... 

The methods of nucleotide synthesis thus far described have involved the activation of a phosphoric moiety by increase of the electrophilic nature of phosphorus, in order to facilitate nucleophilic attack by hydroxyl group(s) of nucleosides (alcoholysis). An alternative approach is the use of an activated nucleoside which will undergo attack by a phosphono-oxy anion on carbon (phosphorolysis). Although the latter approach has had little practical advantage in the synthesis of mononucleotides, it has shown some promise in the synthesis of dinucleoside phosphates. Theoretical considerations are sufficiently compelling to warrant discussion of this approach here. 

The method colmnn denotes procedures or processes used in the synthesis of the nucleotides. The Roman munerals refer to the phos-phorylating agent or system described in Table I. The other abbreviations denote the following AN = by way of anhydronucleotide intermediate(s) D = deamination Enz = enzymic hydrolysis pE] = hydrogenation HA = halogenation Hy = hydrolysis NA = W-alkylation NOx = iV-oxidation Ph = phosphorolysis of the activated nucleoside by the phosphate anion (seep. 369) R = by rearrangement RO = by ring-... 

Figure 9.13. hPNP-catalyzed phosphorolysis of the purine nucleoside. The guanine leaving group and phosphate nucleophile are /ell separated from the oxacarbenium ion, defining a very dissociative TS. 

Glycogen degradation is a phospho-rolysis reaction (breaking of a bond using a phosphate ion as a nucleophile). Enzymes that catalyze phosphorolysis reactions are named phosphorylase. Because more than one type of phosphorylase exists, the substrate usually is included in the name of the enzyme, such as glycogen phosphorylase or purine nucleoside phosphorylase. 

The separation of phosphohydrox5dmino acids by h.p.l.c. has been described. A very sensitive method for the determination of inorganic phosphate has been described which is based on the formation of a Rhodamine B-phosphomolybdate complex, and the continuous enzsmiic removal of inorganic phosphate from reactions has been achieved by means of the nucleoside phosphorylase-catalysed phosphorolysis of nicotinamide riboside, when the product is ribose 1 -phosphate. ... 

Purine nucleoside phosphorylase (PNP) catalyses the phosphorolysis of purine ribonucleosides and 2 -deoxyribo-nucleosides to the purine base and ribose- or 2-deoxyribose-a-1-phosphate. PNP is a key enzyme in the T-cell-mediated immune response. As such, it is an attractive target in a number of therapeutic areas, such as organ transplantation, T-cell-mediated autoimmune disorders and T-cell proliferative diseases. PNP has been the subject of a thorough and successful structure-based drug design study, of which the following gives only a flavour. 

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