Dominika Strzelecka (Presenter)
University of Warsaw
Authorship: Dominika Strzelecka (1) , Sebastian Chmieliñski (2), Sylwia Bednarek (1), Jacek Jemielity (2), Joanna Kowalska (1)
(1) Division of Biophysics, Instytute of Experimental Physics, Faculty of Physics, University of Warsaw, (2) Center of New Tehnologies, University of Warsaw
| Short Abstract Tandem mass spectrometry with electrospray ionization enables to the performance of the controlled fragmentation of molecules. We applied this technique to perform a broad study on the fragmentation of nucleotide derivatives. Identification and structural analysis of these compounds are important because of the crucial role of nucleotides in many biochemical processes. General conclusions regarding nucleotide fragmentation patterns could be drawn from the results. |
Long Abstract
1. Problem
We would like to understand fragmentation pathways of nucleotides and prepare database of MS/MS spectra for various nucleotides
2. Method information
- Sciex API 3200
- Sciex Q TRAP 3200
- Solutions of nucleotides and nucleotide analogs
3. Results
Synthetic analogs of nucleotides and nucleic acids are useful research tools and an emerging class of modern therapeutics. Mass spectrometry is an invaluable tool in identification and quantification of natural nucleotides, nucleotide-based drugs, and their metabolites. However, the fact that many nucleotides and their synthetic analogs are isomeric or isobaric compounds with similar chromatographic mobility, significantly impairs such analyses. We used our in-house collection of nucleotide analogs modified in the phosphate chain, nucleobase, and within ribose moiety to perform a broad study on fragmentation pathways of modified nucleotides. The modifications involved the substitution of hydroxyl groups with various substituents and/or the introduction of the additional groups such as phosphorothioate, phosphoroselenoate, phosphoroamidate, boranophosphate, fluorophosphate, methylenebisphosphonate and imidodiphosphate. In total, we tested over 200 nucleotides with different modifications. We also used isotope labelling with 2H, 18O and 13C and MSn fragmentation to gain insight into fragmentation pathways. Based on the results we proposed general rules for nucleotides fragmentation that may be helpful in MS/MS spectra prediction and identification of unknown compounds based on their fragmentation spectrum. The results of MS/MS were collected in a form of data base which, will be freely available in public domain. The presentation will focus on selected examples demonstrating how MS/MS analyses can be applied to distinguishing between structurally similar isomeric and isobaric nucleotides and to quantification of selected biologically-relevant nucleotides such as mRNA cap metabolites.
4. Conclusions
Based on our research, it can be concluded that tandem mass spectrometry of nucleotides, especially in the negative ion mode, is a valuable source of information on the presence, type and position of modifications. Analysis of fragmentation spectra enables discrimination between isomers and isobars. We hope that the results of our research and the database will be useful to researchers focused on investigation of nucleotide-based drug and prodrug metabolism, discovery of new natural nucleotide modifications in biological samples, as well as chemical synthesis of nucleotide analogs. Development of methods for quantification of mRNA cap metabolites can provide better understanding of degradation pathways of mRNA.
References & Acknowledgements:
| Description | Y/N | Source |
| Grants | no | |
| Salary | no | |
| Board Member | no | |
| Stock | no | |
| Expenses | no |
IP Royalty: no
| Planning to mention or discuss specific products or technology of the company(ies) listed above: | no |