Nucleoside 5 -diphosphate chemical synthesis

The chemical synthesis of a sugar nucleotide diphosphate makes use of an activated nucleotide monophosphate [23] and a glycosyl monophosphate. The most efficient nucleoside monophosphates are phosphorimidazolidates... 

When chemical and enzymatic methods for NTP synthesis are compared[941, enzymatic techniques provide the most convenient route to CTP and GTP, whereas chemical deamination of CTP is the best method for preparing UTP 94L ATP is relatively inexpensive from commercial sources, although it has been synthesized enzymatically from AMP on 50 mmol scale. Mixtures of NTPs can be prepared from RNA by sequential nuclease Pi, polynucleotide phosphorylase, and pyruvate kinase-catalyzed reactions11101. This mixture can be selectively converted to a sugar nucleotide using a particular sugar nucleoside diphosphate pyrophosphorylase11101. 

The chemical synthesis of mono-, di-, tri-, and tetraphosphates usually starts with unprotected nucleosides to produce posphoamidates at first. The monophosphate may be treated either with morpholine and DCC to afford the phosphoro-morpholidate or with carbonyl bis(imidazole) to yield the phosphor imidazolidate. A third method uses diphenylphosphorochloridate to give a pyrophosphate triester (Scheme 8.5.10). All three intermediates react with orthophosphate, diphosphate, or ATP to yield di-, tri-, or tetraphosphates, respectively. 

Whitesides and coworkers have carried out a comparison of enzymic and chemical routes to CTP, GTP and UTP on a 10-gram scale. They concluded that CTP and GTP were best made enzymically, and UTP by reaction of CTP with nitrous acid. The triphosphates were then employed for the enzymic synthesis of UDP-Glucose, UDP-Glucuronic acid, and GDP-Mannose.i94 Cytidine diphosphate sugars have been prepared from the 3,6-dideoxyhexoses paratose and abequose,193 and all four nucleoside diphosphate sugars of 6-sulpho-a-D-quinovose have been synthesized for studies of sulpholipid biosynthesis in chloroplasts.196 The stable analogue (138) of CMP-KDO has been prepared by a triester approach, but was only a weak inhibitor of KDO incorporation into lipopolysaccharides.197 A reference to acetylated forms of UDPGlc is mentioned in Chapter 7. 

GDP-fucose is a glycosyl donor substrate for fucosyltransferases. Although it has been synthesized by many chemical methods [47], the use of fucosyl phosphite in the synthesis seems the most practical [48]. A recent improvement of this method includes the addition of tetrazole to the morpholidate coupling reaction [48b]. This method might also be applicable to the synthesis of other nucleoside diphosphate sugars (Figure 18). 

Nucleoside analogues require phosphorylation to the 5 -triphosphate by cellular kinases prior to their inhibition of DNA synthesis. However, most studies have focused upon prodrugs of monophosphates, whereas for some nucleosides (for example, AZT) the formation of the diphosphate is rate limiting. Huynh-Dinh and coworkers [109, 110] have recently prepared P-acyl esters of 5 -di- (m = I) and tri-phosphates (m = 2) of both AZT and d4T (97, n = 6, 10, 12 or 14, Nu = 5 -AZT or 5 -d4T). The lipophilic acyl chain should facilitate passive diffusion across cellular membranes. Chemical hydrolyses at 37°C and physiological pH showed that the acyl phosphates were cleanly converted to the corresponding nucleoside di- and tri-phosphates. 

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