Electrospray ionization tandem mass spectrometry of biphenyl-modified oligo(deoxy)ribonucleotides

Antisense oligonucleotides and aptamers are important candidates for future therapeutic applications. Different structural modifications are introduced into oligonucleotides to obtain high affinity and binding specificity. Sequence elucidation of oligonucleotides incorporating a wide variety of modifications presents an analytical challenge, as the standard protocols cannot be applied. Mass spectrometry has the potential to solve complex structural problems. However, a better understanding of the fundamental aspects of gas‐phase dissociation of modified DNA and RNA is needed. In this work the influence of specific chemical modifications on backbone dissociation is pointed out. Biphenyl‐modified oligo(deoxy)ribonucleotides, which incorporate C‐glycosidic bound abasic nucleobase substitutes, were subjected to collision‐induced dissociation in an electrospray tandem mass spectrometer, with the goal to investigate the role of nucleobase loss on backbone dissociation. DNA bearing biphenyl nucleobase substitutes show abundant [a‐B]‐ and w‐ions generated by cleavage of the 3′‐CO bonds, except for the phosphodiester groups adjacent to the biphenyl modifications. At these positions no dissociation was observed, demonstrating the dependence of DNA backbone dissociation on nucleobase loss. Also, no evidence for a base loss independent mechanism responsible for formation of w‐ions was found. RNA incorporating biphenyl nucleobase substitutes fragment into c‐ and y‐ions resulting from cleavage of the 5′‐PO bond. Adjacent to the biphenyl modifications no altered dissociation behavior was found. This leads to the conclusion that dissociation of RNA is independent of the 1′‐modification and, therefore, independent of nucleobase loss. Copyright © 2006 John Wiley & Sons, Ltd.