Nucleoside 2 ,3 -phosphates anhydrides

The favored formation of a nucleoside 3 -(alkyl phosphate) was observed on treating the diphenyl phosphate anhydride of uridine 2, 3 -cyclic phosphate with benzyl alcohol.273 In a somewhat related reaction, on treatment with dicyclohexylcarbodiimide in aqueous pyridine, adenosine 2 (3 )-phosphate gave, initially, the 2, 3 -cyclic phosphate, which, on further reaction with the diimide, gave a mixture of the N-phosphonourea nucleoside 44 and its 2 -isomer, in unequal amounts.269 This type of reaction does not seem to occur with... 

Where the method involves cyclization of a nucleoside phosphate to a cyclic phosphate, the following symbols are used for the cyclization reagent DCC = W,W -dicyclohexylcarbodiimide FDNB = l-fluoro-2,4-dinitrobenzene ECF = ethyl chloroformate PC = picryl chloride TFAc = trifluoroacetic acid TFAn = trifluoroacetic anhydride TBO = potassium feri-butoxide in methyl sulfoxide. 

Chapter 3 introduces ATP as a nucleoside triphosphate building block involved in RNA synthesis. The hydrolysis of the terminal phosphate anhydride bond when ATP is converted into adenosine diphosphate (ADP) releases energy. Further hydrolysis of ADP is possible, giving rise to adenosine monophosphate (AMP), with the release of a slightly smaller amount of energy. 

Acetyl phosphate, which can be synthesized from acetic anhydride and phosphoric acid [526], is a commonly used regeneration system in conjunction with acetate kinase [527]. It is modestly stable in aqueous solution and while its phosphoryl donor potential is lower than that of PEP, it is considerably cheaper. As for pymvate kinase, acetate kinase also can accept nucleoside phosphates other than adenosine, and it is inhibited by acetate. 

A primed numeral locates the position of the phosphate on the sugars of mononucleotides (eg, 3 -GMP, 5 -dCMP). Additional phosphoryi groups linked to the first by acid anhydride bonds form nucleoside diphosphates and triphosphates. 

Attention has been drawn to the potential of phosphoric acid anhydrides of nucleoside 5 -carboxylic acids (14) as specific reagents for investigating the binding sites of enzymes. For example, (14 B = adenosine) inactivates adenylosuccinate lyase from E. coli almost completely, but has little effect on rabbit muscle AMP deaminase. The rate of hydrolysis of (14) is considerably faster than that of acetyl phosphate, suggesting intramolecular assistance by the 3 -hydroxyl group or the 3-nitrogen atom. 

Whereas imidazolides of nucleotides react only in organic solvents with phosphates or pyrophosphates to give the corresponding anhydride derivatives in high yield, ATP can also be formed enzymatically in aqueous solution from AMP-Im with inorganic pyrophosphate in the presence of valyl-f-RNA synthetase.[66] A variant of this method is the one-pot reaction of a nucleoside with phosphoryltristriazole and tributylammonium pyrophosphate. 671 An a-methylphosphonyl-/ ,y-diphosphate of a thymidine derivative has been synthesized in a similar way t681... 

Nucleotides are formed when one or more phosphate groups is attached to the 5 carbon of a nucleoside (Figure 1-1-6). Nucleoside di- and triphosphates are high-energy compounds because of the hydrolytic energy associated with the acid anhydride bonds (Figure 1-1-7). 

In ATP, a chain of three phosphate residues is linked to the 5 -OH group of the nucleoside adenosine (see p. 80). These phosphate residues are termed a, p, and y. The a phosphate is bound to ribose by a phosphoric add ester bond. The linkages between the three phosphate residues, on the other hand, involve much more unstable phosphoric add anhydride bonds. The active coenzyme is in fact generally a complex of ATP with an Mg "" ion, which is coordinatively bound to the a and p phosphates (Mg "" ATP ). However, the term ATP is usually used for the sake of simplicity. 

A convenient procedure for synthesis of pyrophosphates by this method was developed by Michelson.317,318 It employs mixed anhydrides (72) formed from nucleoside 5 -phosphates and diphenyl... 

Nucleoside 5 -phosphorothioates have also been employed as activated nucleotide derivatives for synthesis of pyrophosphates.321 The interaction of tributylammonium 2, 3 -di-0-benzoyluridine 5 -phosphorothioate (73) with silver a-D-glucopyranosyl and a-D-galac-topyranosyl phosphates in pyridine solution, with subsequent de-benzoylation, gave the corresponding glycosyl esters in 60-70% yield. This procedure can probably be classified as a variant of the mixed-anhydride method, the driving force of the reaction being the formation of insoluble silver sulfide. 

The energy released by hydrolysis of ATP and the other nucleoside triphosphates is accounted for by the structure of the triphosphate group. The bond between the ribose and the a phosphate is an ester linkage. The a, ft and ft,y linkages are phosphoanhydrides (Fig. 8-40). Hydrolysis of the ester linkage yields about 14 kJ/mol under standard conditions, whereas hydrolysis of each anhydride bond yields about 30 kJ/mol ATP hydrolysis often plays an important thermodynamic role in biosynthesis. When coupled to a reaction with a positive free-energy change, ATP hydrolysis shifts the equilibrium of the overall process to favor product forma-... 

Nucleotides in aqueous solution can be alkylated at the phosphate (and in some cases the nucleoside also) by the action of l-oxidopyridin-2-yldiazo-methane (33). - The protecting group may be removed from the phosphate with snake venom phosphodiesterase, or generally by acetic anhydride treatment, followed by ammonia. Phosphoramidates have been described previously as phosphate-protecting groups, and if 2-naphthylamine is used as its anilidate for this purpose, organic solvent extraction (as above) is possible. A variation on this theme is to use dianilidophosphochloridate (34) as a... 

Adenylic acids o and b were each converted by trifluoroacetic anhydride into the same 2 3 -cyclic phosphate (13, R = adenin-9-yl) by Brown, Magrath, and Todd. These authors also applied this method to the synthesis of 2 3 -cyclic phosphates (13) of cytidine, uridine, and guanosine. Hydrolysis of these cyclic phosphates gave the corresponding a and b nucleotide mixtures. Markham and Smith found that, during hydrolysis of ribonucleic acid with pancreatic ribonuclease, the 2 3 -cyclic phosphates of the pyrimidine nucleosides are formed as intermediates leading to the ribonucleoside phosphates b. The also showed that 2 3 -cyclic phosphates (13) are formed by very mild, alkaline hydrolysis of ribonucleic acid. The discoveries of these a and b isomers of mononucleotides from... 

The nucleotide anhydride, adenosine 5 -triphosphate (24), when digested with aqueous barium hydroxide, gives a complex mixture containing such products as adenine, adenosine, adenosine 2 -, 3 -, and 5 -phosphates, adenosine 5 -pyrophosphate, and adenosine 2 (or 3 ),5 -diphosphate. - In addition, a nucleotide was foimd in this digest whose structure proved - to be that of adenosine 3 5 -cyclic phosphate (25). This component did not consume metaperiodate, and was degraded enzymically to adenosine 5 -phosphate (26) and adenosine 3 -phosphate (27), without the formation of adenosine 2 -phosphate. Hydrolysis of (25) with an acidic ion-exchange resin did, however, produce the 2 - and 3 -phosphates of adenosine. Compound (25) possessed only one phosphoryl dissociation, and showed a ratio of nucleoside to phosphate of 1 1, which, along with a molecular-... 

The role of the APS sulfohydrolases in sulfate metabolism is not understood precisely. The lysosomal APS sulfohydrolase can hydrolyze bis(4-nitro-phenyl) phosphate and 4-nitrophenyl 5 -phosphothymidine (Roger et al., 1978). Thus the lysosomal APS sulfohydrolase is less specific than its cytosolic counterpart, which does not hydrolyze these nitrophenyl derivatives. The apparent role of the lysosomal enzyme is to hydrolyze the acid anhydrides of such compounds as FAD, ATP, and ADP in secondary lysosomes. Thus lysosomal APS sulfohydrolase is an acid anhydride hydrolase that helps the cell in the recovery of nucleoside monophosphates from acid anhydrides. The APS sulfohydrolase in the cytosolic fraction probably regulates the concentrations of PAPS and therefore plays an important role in the control of sulfate conjugation. 

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