Differentiating Positional Isomers of Nucleoside Modifications by Higher-Energy Collisional Dissociation Mass Spectrometry (HCD MS)

Jora, Manasses; Burns, Andrew P.; Ross, Robert L; Lobue, Peter A.; Zhao, Ruoxia; Palumbo, Cody M.; Beal, Peter A.; Addepalli, Balasubrahmanyam; Limbach, Patrick A.

doi:10.1007/s13361-018-1999-6

Cited by 52 publications

(65 citation statements)

References 29 publications

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“…The nucleoside digest was reconstituted in mobile phase A (MPA) (5 mM ammonium acetate, pH 4.5) and separated on a Waters HSS T3 column (100 Å, 1.8 m, 2.1 by 50 mm). Mass spectral data were obtained on a Thermo Scientific Orbitrap Fusion Lumos Tribrid mass spectrometer, following the chromatographic and mass spectrometry conditions previously reported (23).…”

Section: Methodsmentioning

confidence: 99%

“…For example, cnm 5 U has recently been reported in four archaea (H. volcanii, Haloarcula marismortui, Halobacterium salinarum, Methanococcus maripaludis) (22), and we report here the presence of both this position 34 modification and its 2-thiolated analog, cnm 5 s 2 U, which has not been previously characterized. The identification of cnm 5 s 2 U was easily enabled by higher-energy collisional dissociation (HCD) LC-MS/MS of nucleosides (23). Figure 1A shows the HCD MS/MS spectrum obtained when m/z 284, corresponding to cnm 5 U, was dissociated.…”

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confidence: 99%

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tRNA Modification Profiles and Codon-Decoding Strategies in Methanocaldococcus jannaschii

Jora

Solivio

et al. 2019

J Bacteriol

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tRNAs play a critical role in mRNA decoding, and posttranscriptional modifications within tRNAs drive decoding efficiency and accuracy. The types and positions of tRNA modifications in model bacteria have been extensively studied, and tRNA modifications in a few eukaryotic organisms have also been characterized and localized to particular tRNA sequences. However, far less is known regarding tRNA modifications in archaea. While the identities of modifications have been determined for multiple archaeal organisms, Haloferax volcanii is the only organism for which modifications have been extensively localized to specific tRNA sequences. To improve our understanding of archaeal tRNA modification patterns and codondecoding strategies, we have used liquid chromatography and tandem mass spectrometry to characterize and then map posttranscriptional modifications on 34 of the 35 unique tRNA sequences of Methanocaldococcus jannaschii. A new posttranscriptionally modified nucleoside, 5-cyanomethyl-2-thiouridine (cnm 5 s 2 U), was discovered and localized to position 34. Moreover, data consistent with wyosine pathway modifications were obtained beyond the canonical tRNA Phe as is typical for eukaryotes. The high-quality mapping of tRNA anticodon loops enriches our understanding of archaeal tRNA modification profiles and decoding strategies. IMPORTANCE While many posttranscriptional modifications in M. jannaschii tRNAs are also found in bacteria and eukaryotes, several that are unique to archaea were identified. By RNA modification mapping, the modification profiles of M. jannaschii tRNA anticodon loops were characterized, allowing a comparative analysis with H. volcanii modification profiles as well as a general comparison with bacterial and eukaryotic decoding strategies. This general comparison reveals that M. jannaschii, like H. volcanii, follows codon-decoding strategies similar to those used by bacteria, although position 37 appears to be modified to a greater extent than seen in H. volcanii.

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Section: Methodsmentioning

confidence: 99%

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confidence: 99%

tRNA Modification Profiles and Codon-Decoding Strategies in Methanocaldococcus jannaschii

Jora

Solivio

et al. 2019

J Bacteriol

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“…Data were acquired and analyzed using Xcalibur 2.0.7 software (Thermo Fisher Scientific). The option of “all ion fragmentation” using the high-energy collision dissociation (HCD) cell was applied to investigate the confirmation potential of generated fragments and was turned on during the actual analysis [ 46 ].…”

Section: Methodsmentioning

confidence: 99%

Mass Spectrometry-Based Metabolomics Combined with Quantitative Analysis of the Microalgal Diatom (Chaetoceros calcitrans)

et al. 2020

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Although many metabolomics studies of higher land plant species have been conducted, similar studies of lower nonland plant species, which include microalgae, are still developing. The present study represents an attempt to characterize the metabolic profile of a microalgal diatom Chaetoceros calcitrans, by applying high-resolution mass spectrometry detection, via Q-ExactiveTM Plus Orbitrap mass spectrometry. The results showed that 54 metabolites of various classes were tentatively identified. Experimentally, the chloroform and acetone extracts were clearly distinguished from other solvent extracts in chemometric regression analysis using PLS, showing the differences in the C. calcitrans metabolome between the groups. In addition, specific metabolites were evaluated, which supported the finding of antioxidant and anti-inflammatory activities. This study also provides data on the quantitative analysis of four carotenoids based on the identification results. Therefore, these findings could serve as a reliable tool for identifying and quantifying the metabolome that could reflect the metabolic activities of C. calcitrans.

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“…The comparison of our recorded spectra to previously published data 30 revealed an additional variation in the fragmentation patterns (Supplementary Figure S3 Figure S5C). Similar fingerprint ionization patterns for distinction of isomers has been characterized before using higher-energy collisional dissociation MS 35 .…”

Section: Discussionmentioning

confidence: 76%

Broad-range RNA modification analysis of complex biological samples using rapid C18-UPLC-MS

Gregorová

Sipari

Sarin

2020

Preprint

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Post-transcriptional RNA modifications play an important role in cellular metabolism with homeostatic disturbances manifesting as a wide repertoire of phenotypes, reduced stress tolerance and translational perturbation, developmental defects, and diseases, such as type II diabetes, leukemia and carcinomas. Hence, there has been an intense effort to develop various methods for investigating RNA modifications and their roles in various organisms, including sequencing-based approaches and, more frequently, liquid chromatography mass spectrometry (LC-MS)-based methods. Although LC-MS offers numerous advantages, such as being highly sensitive and quantitative over a broad detection range, some stationary phase chemistries struggle to resolve positional isomers. Furthermore, the demand for detailed analyses of complex biological samples often necessitates long separation times, hampering sample-to-sample turnover and making multisample analyses time consuming. To overcome this limitation, we have developed an ultra-performance LC-MS (UPLC-MS) method that uses an octadecyl carbon chain (C18)-bonded silica matrix for the efficient separation of 50 modified ribonucleosides, including positional isomers, in a single 9 min sample-to-sample run. To validate the performance and versatility of our method, we analyzed tRNA modification patterns of representative microorganisms from each kingdom of life, namely Archaea (Methanosarcina acetivorans), Bacteria (Pseudomonas syringae) and Eukarya (Saccharomyces cerevisiae). Additionally, our method is flexible and readily applicable for detection and relative quantification using stable isotope labelling and targeted approaches like multiple reaction monitoring (MRM). In conclusion, this method represents a fast and robust tool for broad-range exploration and quantification of ribonucleosides, facilitating future homeostasis studies of RNA modification in complex biological samples.

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Differentiating Positional Isomers of Nucleoside Modifications by Higher-Energy Collisional Dissociation Mass Spectrometry (HCD MS)

Cited by 52 publications

References 29 publications

tRNA Modification Profiles and Codon-Decoding Strategies in Methanocaldococcus jannaschii

tRNA Modification Profiles and Codon-Decoding Strategies in Methanocaldococcus jannaschii

Mass Spectrometry-Based Metabolomics Combined with Quantitative Analysis of the Microalgal Diatom (Chaetoceros calcitrans)

Broad-range RNA modification analysis of complex biological samples using rapid C18-UPLC-MS

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