Abstract
With the development of new synthesis procedures, an ever increasing number of chemical modifications can now be incorporated into synthetic oligonucleotides, representing new challenges for analytical chemists to efficiently identify and characterize such molecules. While conventional mass spectrometry (MS) has proven to be a powerful tool to study nucleic acids, new and improved methods and software are now needed to address this emerging challenge. In this report, we describe a simple yet powerful program that affords great flexibility in the calculation of theoretical masses for conventional as well as modified oligonucleotide molecules. This easy to use program can accept input oligonucleotide sequences and then calculate the theoretical mass values for full length products, process impurities, potential metabolites, and gas phase fragments. We intentionally designed this software so that modified nucleotide residues can be incorporated into oligonucleotide sequences, and corresponding mass values can be rapidly calculated. To test the utility of this program, two oligonucleotides that contain a large number of chemical modifications were synthesized. We have analyzed these samples using a Q-TOF mass spectrometer and compared the calculated masses to the observed ones. We found that all of the data matched very well with less than 30 ppm mass errors, well within the expectation for our instrument operated in its current mode. These data confirmed the validity of calculations performed with this new software.
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References
Bennett, C.F., Swayze, E.E.: RNA targeting therapeutics: molecular mechanisms of antisense oligonucleotides as a therapeutic platform. Annu. Rev. Pharmacol. Toxicol. 50, 259–293 (2010)
Keefe, A.D., Pai, S., Ellington, A.: Aptamers as therapeutics. Nat. Rev. Drug Discov. 9(7), 537–550 (2010)
Castanotto, D., Rossi, J.J.: The promises and pitfalls of RNA-interference-based therapeutics. Nature 457(7228), 426–433 (2009)
Gragoudas, E.S., Adamis, A.P., Cunningham, E.T., Feinsod, M., Guyer, D.R.: Pegaptanib for neovascular age-related macular degeneration. New Engl. J. Med. 351(27), 2805–2816 (2004)
Fabris, D.: A role for the MS analysis of nucleic acids in the post-genomics age. J. Am. Soc. Mass Spectrom. 21(1), 1–13 (2010)
Gardner, M.W., Li, N., Ellington, A.D., Brodbelt, J.S.: Infrared multiphoton dissociation of small-interfering RNA anions and cations. J. Am. Soc. Mass Spectrom. 21(4), 580–591 (2010)
Wu, J., McLuckey, S.A.: Gas-phase fragmentation of oligonucleotide ions. Int. J. Mass Spectrom. 237(2/3), 197–241 (2004)
Gao, H., Liu, Y., Rumley, M., Yuan, H.M., Mao, B.: Sequence confirmation of chemically modified RNAs using exonuclease digestion and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Rapid Commun. Mass Spectrom. 23(21), 3423–3430 (2009)
Nikcevic, I., Wyrzykiewicz, T.K., Limbach, P.A.: Detecting low-level synthesis impurities in modified phosphorothioate oligonucleotides using liquid chromatography-high resolution mass spectrometry. Int. J. Mass Spectrom. 304(2/3), 98–104 (2011)
Zou, Y., Tiller, P., Chen, I.W., Beverly, M., Hochman, J.: Metabolite identification of small interfering RNA duplex by high-resolution accurate mass spectrometry. Rapid Commun. Mass Spectrom. 22(12), 1871–1881 (2008)
Available at: http://rna-mdb.cas.albany.edu/RNAmods/masspec/mongo.htm. Accessed 10 April 2013
Rozenski, J., Crain, P.F., McCloskey, J.A.: The RNA Modification Database: 1999 update. Nucleic Acids Res. 27(1), 196–197 (1999)
Verma, S., Eckstein, F.: Modified oligonucleotides: synthesis and strategy for users. Annu. Rev. Biochem. 67, 99–134 (1998)
Rozenski, J., McCloskey, J.A.: SOS: a simple interactive program for ab initio oligonucleotide sequencing by mass spectrometry. J. Am. Soc. Mass Spectrom. 13(3), 200–203 (2002)
Available at: http://www.enovatia.com/products/promass/. Accessed 10 April 2013
Acknowledgments
The authors especially acknowledge Gary Martin and Chris Welch for their helpful comments on this manuscript. The authors thank Huimin Yuan, Vinny Antonucci, and Zhihong Ge for their managerial support. In addition, we thank Jef Rozenski for access of the SOS software and helpful discussions.
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Yang, J., Leopold, P., Helmy, R. et al. Design and Application of an Easy to Use Oligonucleotide Mass Calculation Program. J. Am. Soc. Mass Spectrom. 24, 1315–1318 (2013). https://doi.org/10.1007/s13361-013-0643-8
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DOI: https://doi.org/10.1007/s13361-013-0643-8