Tandem mass spectrometric sequence characterization of synthetic thymidine‐rich oligonucleotides

Abdullah, A. M.; Sommers, Cynthia D.; Hawes, Jessica J.; Rodriguez, Jason D.; Yang, Kui

doi:10.1002/jms.4819

Cited by 10 publications

(10 citation statements)

References 30 publications

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“…The sequence of the nucleotide chain must be confirmed. For oligos smaller than approximately 50 bases, tandem mass spectrometry has proven to be the method of choice [ 14 , 15 , 16 ]. Longer ribonucleic acid (RNA) molecules have been successfully sequenced by enzymatic hydrolysis approaches [ 17 ].…”

Section: Introductionmentioning

confidence: 99%

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Investigation of the Influence of Charge State and Collision Energy on Oligonucleotide Fragmentation by Tandem Mass Spectrometry

Gawlig¹,

Rühl²

2023

Molecules

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Due to the increasing pharmaceutical interest of oligonucleotides, for example in antisense therapy and vaccines, their analytical characterization is of fundamental importance due to their complex structure. For this purpose, mass spectrometry is a viable tool for structural studies of nucleic acids. Structural information regarding the primary sequence of a nucleic acid can reliably be gained via tandem mass spectrometry (MSMS) fragmentation. In this work, we present the characteristic fragmentation behavior of short-chain oligonucleotides (15–35 nucleotides) with respect to the collision-induced dissociation (CID) voltage used. The relationship and influence of the length of the oligonucleotide and its charge state is also discussed. The results presented here can be helpful for estimating the required fragmentation energies of short-chain oligonucleotides and their sequencing.

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Section: Introductionmentioning

confidence: 99%

“…With this method, it is possible to gain a unique dissociation pattern, and thus a structural fingerprint of the analyte. Through high measurement resolution and the determination of a precursor ion, the occurrence of foreign fragments can be significantly reduced and the elucidation thus simplified considerably [ 14 , 16 ].…”

Section: Introductionmentioning

confidence: 99%

Investigation of the Influence of Charge State and Collision Energy on Oligonucleotide Fragmentation by Tandem Mass Spectrometry

Gawlig¹,

Rühl²

2023

Molecules

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“…Abdullah et al 74 following supplemental collisional activation. The addition of Co adds more complexity through further salt adduction observed in the mass spectra, making manual processing more time consuming.…”

Section: Other Ms/ms Techniques To Characterise Oligonucleotidesmentioning

confidence: 99%

Review of fragmentation of synthetic single‐stranded oligonucleotides by tandem mass spectrometry from 2014 to 2022

Black

Ray

Stulz

et al. 2023

Rapid Comm Mass Spectrometry

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The fragmentation of oligonucleotides by mass spectrometry allows for the determination of their sequences. It is necessary to understand how oligonucleotides dissociate in the gas phase, which allows interpretation of data to obtain sequence information. Since 2014, a range of fragmentation mechanisms, including a novel internal rearrangement, have been proposed using different ion dissociation techniques. The recent publications have focused on the fragmentation of modified oligonucleotides such as locked nucleic acids, modified nucleobases (methylated, spacer, nebularine and aminopurine) and modification to the carbon 2′‐position on the sugar ring; these modified oligonucleotides are of great interest as therapeutics. Comparisons of different dissociation techniques have been reported, including novel approaches such as plasma electron detachment dissociation and radical transfer dissociation. This review covers the period 2014–2022 and details the new knowledge gained with respect to oligonucleotide dissociation using tandem mass spectrometry (without priori sample digestion) during that time, with a specific focus on synthetic single‐stranded oligonucleotides.

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“…Most of the early investigations concentrated on the dissociation mechanisms of oligonucleotides that contained one to three nucleotides (nt). Many fragmentation pathways were discovered from analyses of DNA and RNA, including a variety of sequence-determining ions from backbone cleavage, neutral or charged nucleobase losses from precursor ions, and released charged-nucleotides, [30][31][32][33][34][35][36][37][38] but these published discrepant data remains a bit turbid for oligonucleotide fragmentation. 30 Additionally, far fewer mechanistic studies have addressed RNA dissociation than DNA dissociation.…”

Section: Introductionmentioning

confidence: 99%

Charge-State-Dependent Collision-Induced Dissociation Behaviors of RNA Oligonucleotides via High-Resolution Mass Spectrometry

Sun

Zuo

Zhang

et al. 2023

Preprint

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Mass spectrometry (MS)-based analysis of RNA oligonucleotides (oligos) plays an increasingly important role in the development of RNA therapeutics and in epitranscriptomic studies. However, MS fragmentation behaviors of RNA oligos are understood insufficiently. In this study, we characterized the negative-ion-mode fragmentation behaviors of 26 synthetic RNA oligos of four to eight nucleotides (nt) in length by collision-induced dissociation (CID) using a high-resolution, accurate-mass instrument. We find that in the CID spectra acquired under the normalized collisional energy of 35%, ~70% of the total peak intensity belonged to sequencing ions (a-B, a, b, c, d, w, x, y, z), ~25% belonged to precursor ions with either complete or partial loss of a nucleobase in the form of a neutral or an anion, and the remainder were internal ions and anionic nucleobases. Of the sequencing ions, the most abundant species were y, c, w, a-B, and a ions. The charge state of the RNA precursor ions strongly affected their fragmentation behaviors. As the precursor charge increased from -1 to -5, the fractional intensity of sequencing ions in the CID spectra decreased, whereas the fractional intensity of precursor ions with neutral and/or charged losses of a nucleobase increased. Moreover, RNA oligos containing U, especially at the 3′ terminus, tended to produce precursors that lost HNCO and/or NCO-, which presumably corresponded to isocyanic acid and cyanate anion, respectively. These findings build a strong foundation for mechanistic understanding of RNA fragmentation by MS/MS, contributing to future automated identification of RNA oligos from their CID spectra in a more efficient way.

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Tandem mass spectrometric sequence characterization of synthetic thymidine‐rich oligonucleotides

Cited by 10 publications

References 30 publications

Investigation of the Influence of Charge State and Collision Energy on Oligonucleotide Fragmentation by Tandem Mass Spectrometry

Investigation of the Influence of Charge State and Collision Energy on Oligonucleotide Fragmentation by Tandem Mass Spectrometry

Review of fragmentation of synthetic single‐stranded oligonucleotides by tandem mass spectrometry from 2014 to 2022

Charge-State-Dependent Collision-Induced Dissociation Behaviors of RNA Oligonucleotides via High-Resolution Mass Spectrometry

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