Potential Pitfalls in MALDI-TOF MS Analysis of Abiotically Synthesized RNA Oligonucleotides

Burcar, Bradley T.; Cassidy, Lauren M.; Moriarty, Elizabeth M; Joshi, P. C.; Coari, Kristin M.; McGown, Linda B.

doi:10.1007/s11084-013-9334-5

Cited by 28 publications

(38 citation statements)

References 30 publications

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“…For shorter polymers, we also obtained data from ESI‐MS‐calibrated MALDI‐TOF measurements, which are ideal to resolve lengths of fewer than ten nucleotides. As both techniques are prone to show false‐positive polymerization signals due to stacking of nucleotides (particularly pronounced with purine bases),15 we also performed digestion by RNAse T1, which is specific to 3′–5′‐linked RNA strands.…”

Section: Resultsmentioning

confidence: 99%

Dry Polymerization of 3′,5′‐Cyclic GMP to Long Strands of RNA

et al. 2014

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Recent progress in the synthesis of nucleotides from prebiotically plausible precursors has opened up new ways to explain the origin of genetic matter. Mechanisms for the polymerization of nucleotides without the help of catalysts are, however, rare. Complementary to the experiments done by Costanzo et al., we found that drying 3',5'-cyclic GMP leads to poly-G RNA strands with lengths of up to 40 nucleotides. We also show that the polymerization to long RNA strands is considerably more efficient under dry conditions than for cGMP polymerization in water. The length depends on the incubation time of dry nucleotides at temperatures of 40-80 °C. No enzymes or other catalysts are needed for successful polymerization.

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Section: Resultsmentioning

confidence: 99%

Dry Polymerization of 3′,5′‐Cyclic GMP to Long Strands of RNA

et al. 2014

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“…These signals are displayed in the corresponding blow-ups shown in Fig 3, panels A, B and C. These molecules necessarily derive from a covalent dimer, and are present in the standard and in the fragmentation profiles of both the dimers and trimers. The fragmentation of the trimer (panel C) yields, in addition to these two unambiguous indicators, also a dimer (see the signal corresponding to pApA at m/z = 675.1), which is compatible both with a covalent dimer (as expected from the fragmentation of a trimer) and with a non-covalent adduct of a cAMP and an AMP [12] and is thus considered not resolutory. In conclusion, in spite of the particular tendency of this system to create false-positives, the fragmentation analysis provides direct evidence of the presence of covalently-bound dimers and trimers.…”

Section: Methodsmentioning

confidence: 99%

Non-Enzymatic Oligomerization of 3’, 5’ Cyclic AMP

et al. 2016

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Recent studies illustrate that short oligonucleotide sequences can be easily produced from nucleotide precursors in a template-free non-enzymatic way under dehydrating conditions, i.e. using essentially dry materials. Here we report that 3’,5’ cyclic AMP may also serve as a substrate of the reaction, which proceeds under moderate conditions yet with a lower efficiency than the previously reported oligomerization of 3’,5’ cyclic GMP. Optimally the oligomerization requires (i) a temperature of 80°C, (ii) a neutral to alkaline environment and (iii) a time on the order of weeks. Differences in the yield and required reaction conditions of the oligomerizations utilizing 3’,5’ cGMP and cAMP are discussed in terms of the crystal structures of the compounds. Polymerization of 3’,5’ cyclic nucleotides, whose paramount relevance in a prebiotic chemistry context has been widely accepted for decades, supports the possibility that the origin of extant genetic materials might have followed a direct uninterrupted path since its very beginning, starting from non-elaborately pre-activated monomer compounds and simple reactions.

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“…In general, over the course of these experiments we observed a tendency to detect longer polymers, on average, for the purines relative to the pyrimidines. Peaks at –m/z 18 from linear polymer peaks are attributed to circular polymers that lose a water molecule upon condensation of the linear strand to form the circular Peaks at +m/z 18 or multiples of 18 are attributed to aggregates of linear polymers or aggregates of linear and circular polymer (Burcar et al 2013). …”

Section: Resultsmentioning

confidence: 99%

“…Lower m/z are complicated mixtures of monomer and dimer peaks, MALDI matrix peaks and other small molecular species that make assignments difficult. Peak assignment was performed as previously described (Burcar et al 2013). The LMW oligonucleotide standard was used as an external calibrant and was run prior to all analyses.…”

Section: Methodsmentioning

confidence: 99%

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Nucleotide Selectivity in Abiotic RNA Polymerization Reactions

Coari

Martin

Jain

et al. 2017

Orig Life Evol Biosph

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In order to establish an RNA world on early Earth, the nucleotides must form polymers through chemical rather than biochemical reactions. The polymerization products must be long enough to perform catalytic functions, including self-replication, and to preserve genetic information. These functions depend not only on the length of the polymers, but also on their sequences. To date, studies of abiotic RNA polymerization generally have focused on routes to polymerization of a single nucleotide and lengths of the homopolymer products. Less work has been done the selectivity of the reaction toward incorporation of some nucleotides over others in nucleotide mixtures. Such information is an essential step toward understanding the chemical evolution of RNA. To address this question, in the present work RNA polymerization reactions were performed in the presence of montmorillonite clay catalyst. The nucleotides included the monophosphates of adenosine, cytosine, guanosine, uridine and inosine. Experiments included reactions of mixtures of an imidazole-activated nucleotide (ImpX) with one or more unactivated nucleotides (XMP), of two or more ImpX, and of XMP that were activated in situ in the polymerization reaction itself. The reaction products were analyzed using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) to identify the lengths and nucleotide compositions of the polymerization products. The results show that the extent of polymerization, the degree of heteropolymerization vs. homopolymerization, and the composition of the polymeric products all vary among the different nucleotides and depend upon which nucleotides and how many different nucleotides are present in the mixture.

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Potential Pitfalls in MALDI-TOF MS Analysis of Abiotically Synthesized RNA Oligonucleotides

Cited by 28 publications

References 30 publications

Dry Polymerization of 3′,5′‐Cyclic GMP to Long Strands of RNA

Dry Polymerization of 3′,5′‐Cyclic GMP to Long Strands of RNA

Non-Enzymatic Oligomerization of 3’, 5’ Cyclic AMP

Nucleotide Selectivity in Abiotic RNA Polymerization Reactions

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