Nucleosides linkage

In view of the difficulty of hydrolyzing the pyrimidine nucleosidic linkages, ribonucleic acids have been hydrolyzed to a mixture of purine bases and pyrimidine nucleotides which is then separated by paper chromatography.132, 163 164 This method has been employed extensively for the analysis of ribonucleic acids, and gives reproducible results,166 but it has not been used to any great extent for deoxyribonucleic acids, probably because, under these conditions of hydrolysis, they yield some pyrimidine deoxy-ribonucleoside diphosphates.166... 

The nucleoside linkage is formed from the purine N9 or pyrimidine Nj to the Cl of the ribose sugar, and the common names are given below ... 

Generally, two different strategies are used in conventional synthesis of oligonucleotides the diester and the triester method [131,132]. In syntheses with soluble polymer supports and in recent works in general the diester method is preferred. Thereby, the 5 -0 position is protected by the solutde polymer and the elongation of the oligonucleotide chain is carried out via the 3 -0 position. Most researchers used an ether, ester, or a carbonate bond for the S -nucleoside linkage. 

Fig. 5. Solid-phase supports made of synthetic resins. Figs 1 and 2 The nucleoside linkages used on the early polystyrene popcorn supports (3,4). Fig. 3. The nucleoside linkage used on a polyamide based resin (5). Fig. 4 The structure of a polyamide/silica gel composite developed for continuous-flow synthesis (8) Fig. 5. The structure of a PEG-polystyrene grafted copolymer, tentacle support (25). The mean mol wt of die PEG side chain is 3000 daltons. Fig 6 The nucleoside linkage used on the recently introduced highly crosslinked polystyrene support (27).

Fig. 7. Hydrolysis resistant nucleoside linkages. Fig. 1. The TDIC linkage based on tolylene-2,6-diisocyanate (32). Fig. 2 A ribonucleotide linked to LCAA-CPG via a secondary amino group (35) Fig 3. Nucleoside attachment via a 1,6-bis-methylaminohexane linker (36),...

Fig. 8. Easily cleavable nucleoside linkages. Fig 1 A disulfide linkage that yields 3 -phosphorylated products (39) Figs. 2. and 3. Linkages containing a 2-(2-nitrophenyl)ethyl linker, which can be cleaved by DBU, for the synthesis of oligonucleotides with either free or phosphorylated 3 -ends (40) Fig. 4. The oxalyl-CPG linkage (41), which is the most easily hydrolyzable linkage avmlable.

The naturally occurring C-nucleosides containing C-glycosyl linkages are shown in Table 1. 

The 3, 5 intemucleotide linkage is formed either by condensing die 3 -hydroxy group of an appropriately protected deoxyribonucleotide or -nucleoside with the 5 -phosphate of a deoxyribonucleotide (method a), or by condensing a 3 -phosphordiester with the 5 -hydroxy group of a nucleoside in a modified phosphortriester approach (method b). 

Imidazolides of adenylic acid (ImpA) or uridylic acid (ImpU) are polycondensed to oligonucleotides by means of Zn2+ ions. 1673 The resulting phosphordiester bond was found to be of the 2, 5 type. In the reaction of nucleoside 5 -phosphoric acid methyl ester with ImpA in the presence of MgC, 2, 5 -dinucleotides are formed six to nine times more frequently than the corresponding 3, 5 compounds. 63 Polycondensations of ImpA in aqueous solution in the presence of various divalent metal ions lead to short oligo-adenylic acids (pA) (n = 1—5) mainly with 2, 5 -intemucleotide linkages. With Pb2+, for example, the total yield of oligomers was as high as 57%. 1683 1693... 

The problems in the nucleoside synthesis arise in the linkage of the 3-N atoms of the pyrimidines and the 9-N atoms of the purines with the l -C atom of ribose, not only without enzyme control, but also under conditions extant on the primordial Earth. How might such reactions occur There have naturally been many attempts... 

The position was somewhat clarified by the isolation of 2- and 3-O-phos-phonucleosides from ribonucleic acid hydrolyzates in 92 to 100% yields,134 and also by the demonstration that 5-O-phosphonucleosides are also present in enzymic digests.49, 197 Yet this information gave no indication of the nature of the alkali-labile linkages. Thus, while the majority of the experimental evidence pointed to the phosphoryl residues as being doubly esterified with adjacent nucleosides, two facts remained apparently inexplicable on the basis of this type of structure. First, ready fission by alkalis, and secondly, the absence of 5-phosphates from alkaline hydrolyzates and their presence in enzymic digests. Both these facts have been explained by Brown and Todd in the following way.92... 

Hydrolysis of ribonucleic acids by snake venom was found to yield inorganic phosphate, nucleosides, and pyrimidine ribonucleoside diphosphates.197 These diphosphates were shown by their behavior toward various enzymes to be mixtures of 2,5- and 3,5-diphosphates, and it therefore seems likely that they were formed through intermediate, cyclic phosphates. Thus, although this evidence confirms the existence of 2(or 3) — 5 linkages, it does not distinguish between the 2- and 3-positions. 

The most convincing evidence in favor of a uniform 3,5-diester linkage between nucleotides has been obtained by the action of various enzymes on synthetic diesters of known constitution.218 217 Ribonuclease and spleen extracts were found to act only on nucleoside 3-(benzyl hydrogen phosphates), but not on other isomers, to give nucleoside cyclic phosphates which are broken down further to give nucleoside 3-phosphates. It is concluded, by analogy, that polynucleotides, which are substrates for these enzymes, also possess ester groupings at the 3-positions, rather than at the... 

The major missing component for the formation of RNA as an information-bearing molecule is the distinct lack of ribose in any particularly large quantities. The backbone of RNA and DNA has the phosphoester linkage linking the nucleosides - the bases linked with ribose, as seen in Figure 8.10a. 

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