Letsinger-Caruthers synthesis

SYNTHESIS OF DNA The last section of this report concerns the Letsinger-Caruthers synthesis of DNA. It seems to me that an introductory paper should concentrate on the most important topics, even if my own contribution to the topic is pretty small. Biochemists today isolate and replicate the genes for many enzymes, and then express and isolate proteins related to those genes. They can work up a piece of nucleic add from a tiny sample — a few billion molecules — to make milligrams of nucleic acid and eventually express many milligrams of enzyme. 

The Letsinger-Caruthers synthesis is unrelated to the biochemical one except in the area of efficiency. Because phosphites are so reactive, they would be totally unsuitable for the genetic tape itself, but because they are so reactive, they are ideally suited as intermediates in the chemical synthesis of polynucleotides. One can accomplish the needed condensation reactions with phosphites under strictly anhydrous conditions with very weakly acidic catalysts, and then stabilize the product by a mild oxidation of the phosphite to the desired phosphate with iodine. 

Fig. 9. The Letsinger - Caruthers synthesis of a protected form of nucleotides.

Although Letsinger s original phosphite synthesis is an efficient process, it was still too difficult for general use, as it required dry ice temperatures and strictly anhydrous conditions. The method, however, was modified by Marvin Caruthers (27) who substituted monoester diamidites for phosphites. The reactions proceed much more smoothly with the amidites, which are, relatively, resistant to hydrolysis and to autoxidation. A beginning of the physical organic chemistry of the phosphites has been published (22), but to the best of my knowledge none has been published for the phosphoramidites. Perhaps this review will stimulate someone to take up this problem,just as Todd s lectures stimulated me to work in this area. 

The development of deoxyribonucleoside phosphoramidite derivatives for the synthesis of oligodeoxyribonucleotides was first described by Beaucage and Caruthers (1) in 1981. The conceptual basis of this methodology emerged as a modification of the phos-phorodichloridite coupling procedure reported earlier by Letsinger and CO workers (2). Essentially, the approach involved the reaction of the protected deoxyribonucleosides la-d (Fig. 1) withchloro-(A(iV-dimethyl-amino) methoxyphosphine (2) in the presence of iV,iV-diiso-propylethylamine. The rapid reaction yielded the deoxyribonucleoside phosphoramidites 3a-d (Fig. 1), which were isolated by precipitation and stored as dry powders. 

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