Nucleoside 3 ,5 -cyclic dinucleoside phosphates

Synthesis by Transfer. Ribonuclease catalyzes transfer reactions in addition to hydrolyses. Both cyclic nucleotides and diesters of 3 -nucleo-tides serve as substrates for the transfer. Acceptors for the phosphate include the primary alcohol groups of nucleosides, nucleoside cyclic phosphates, and simple alcohols. Simple nucleotides do not serve as acceptors. The products of transfer reactions, including dinucleoside monophosphates and the so-called cyclic dinucleotides, can also serve as acceptors, and by repeated transfer reactions of cyclic nucleotides small polymers have been built up (IV). It has been suggested that this reaction may play a role in nucleic acid biosynthesis. ... 

The base specificity has been extensively investigated by the 3 -terminal analysis of digestion products of RNA and polyribonucleotides and by the studies on the susceptibility of dinucleoside monophosphates or nucleoside 2, 3 -cyclic phosphates to the enzyme. The results are summarized in Table V (24-87). From the base specificity study, the es-... 

The observation that the final digestion products of RNA by the enzyme are four nucleoside 2, 3 -cyclic phosphates led Ukita and coworkers (121) to utilize the enzyme for the synthesis of various dinucleoside monophosphates. Indeed they have successfully synthesized UpU, CpU, ApU, GpU, UpC, CpC, ApC, and GpC in good yields (from 20 to 75% based on initial nucleoside 2, 3 -cyclic phosphates) by the reaction... 

In the 1,3,2-dioxaphosphole method a bis(2-butene-2,3-diyl) pyrophosphate is used as the condensing agent. It allows two successive esterifications of one phosphate group to be performed without additional activation. First a 5 -O-protected nucleoside is added in methylene chloride in the second reaction an unprotected nucleoside can be used, since only the 3 OH group is able to attack the cyclic enediol 3 -nucleosidyl phosphotriester. Protected dinucleoside triesters are obtained in 80% yield. Removals of protective groups, methoxytrityl by means of trifluoroacetic acid in methylene chloride and 1-methylacetonyl by aqueous triethyl-amine, also give about 80% yield (F. Ramirez, 1975, 1977). 

The dinucleoside dipyrophosphate 81 has been obtained in 30% yield following reaction of the nucleosides with a large excess of phosphoryl chloride in trimethylphosphate/DMF. Slow decomposition of 81 produces the corresponding nucleoside 3, 5 -cyclic phosphate derivatives. 

Fig. 9-7 A schematic mechanism for the hydrolysis of a dinucleoside by ribonuclease A. The various conformational states inferred to be present from kinetic studies are shown. In this mechanism PypN is a pyrimidine 3 5 -nucleoside, Py 2 3 p is a pyrimidine 2 3 cyclic phosphate and E is the enzyme. The primes designate different conformational states of the enzyme and enzyme-substrate complexes.

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