Synthesis of a 4‐Selenothymidine Phosphoramidite and Incorporation into Oligonucleotides

Abstract

The detailed synthetic protocol for a 4-selenothymidine phosphoramidite and its use to prepare modified oligonucleotides is described here. The Se-phosphoramidite synthesis was achieved by developing a useful protection and deprotection system for the selenium functionality. The coupling reaction of the Se-phosphoramidite during solid-phase oligonucleotide synthesis is quantitative, and the oligonucleotides containing the Se-modification are stable. Based on crystal structure analysis, the selenium-modified oligonucleotides retain base-pairing like their native counterparts, and the derivatized DNA structure is virtually identical to the native structure. This achievement will present a novel opportunity for structural studies of nucleic acids and their protein complexes, because selenium can resolve the phase problem in macromolecular X-ray crystallography. In addition, this atom-specific replacement of oxygen with selenium will provide a useful tool for investigating biochemical and biophysical properties of nucleic acids and their protein complexes.