5(4H)-Oxazolones as Effective Aminoacylation Reagents for the 3′-Terminus of RNA

Liu, Ziwei; Hanson, Cassandra; Ajram, Ghinwa; Boiteau, Laurent; Rossi, Jean‐Christophe; Danger, Grégoire; Pascal, Robert

doi:10.1055/s-0036-1588647

Cited by 8 publications

(8 citation statements)

References 13 publications

(17 reference statements)

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“…In addition, previous work on oxazolone modification of small RNA oligonucleotide models indicates that the vicinal diol and terminal phosphates (2′, 3′, or 5′) are strongly preferred as nucleophiles, with no detectable reactivity at internal 2′-OH sites. 43,44 In contrast, we found that all ribozymes tested, representing each motif (1A, 1B, 2, 3), were modified at an internal 2′-OH. Therefore, the true catalytic enhancement provided by these ribozymes at a specific internal 2′-OH is likely to be at least 700-fold greater (Supporting Text S2) than the r s as reported here.…”

Section: Resultsmentioning

confidence: 66%

“…These sites stand in contrast to the modification of modern tRNAs at the vicinal diol (3′ terminus), which is also found to be more reactive in model oligonucleotides. 43,44 It is possible that an internal reaction site facilitates establishment of multiple contacts with BYO, and the rate acceleration caused by these structural features outweighs the intrinsic reactivity of the vicinal diol. Similarly, it is unknown whether the identity of the 3′ terminal sequence (CUG in this study, compared to CCA in tRNAs) may contribute to this finding.…”

Section: Discussionmentioning

confidence: 99%

“…These compounds react with amino acids to form peptides, 39 and therefore have been proposed as a prebiotic form of chemically activated amino acids. At high concentration, NCAs and 5(4 H )-oxazolones react with phosphate esters, including nucleotides, to form aminoacyl-RNA mixed anhydrides in low yield, 40−44 suggesting this reaction as a candidate for ribozyme catalysis. Use of 5(4 H )-oxazolones avoids uncontrolled polymerization in comparison to NCAs, making oxazolones a practical and prebiotically relevant substrate for in vitro selection.…”

Section: Introductionmentioning

confidence: 99%

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Mapping a Systematic Ribozyme Fitness Landscape Reveals a Frustrated Evolutionary Network for Self-Aminoacylating RNA

et al. 2019

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Molecular evolution can be conceptualized as a walk over a “fitness landscape”, or the function of fitness (e.g., catalytic activity) over the space of all possible sequences. Understanding evolution requires knowing the structure of the fitness landscape and identifying the viable evolutionary pathways through the landscape. However, the fitness landscape for any catalytic biomolecule is largely unknown. The evolution of catalytic RNA is of special interest because RNA is believed to have been foundational to early life. In particular, an essential activity leading to the genetic code would be the reaction of ribozymes with activated amino acids, such as 5(4H)-oxazolones, to form aminoacyl-RNA. Here we combine in vitro selection with a massively parallel kinetic assay to map a fitness landscape for self-aminoacylating RNA, with nearly complete coverage of sequence space in a central 21-nucleotide region. The method (SCAPE: sequencing to measure catalytic activity paired with in vitro evolution) shows that the landscape contains three major ribozyme families (landscape peaks). An analysis of evolutionary pathways shows that, while local optimization within a ribozyme family would be possible, optimization of activity over the entire landscape would be frustrated by large valleys of low activity. The sequence motifs associated with each peak represent different solutions to the problem of catalysis, so the inability to traverse the landscape globally corresponds to an inability to restructure the ribozyme without losing activity. The frustrated nature of the evolutionary network suggests that chance emergence of a ribozyme motif would be more important than optimization by natural selection.

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Section: Resultsmentioning

confidence: 66%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Mapping a Systematic Ribozyme Fitness Landscape Reveals a Frustrated Evolutionary Network for Self-Aminoacylating RNA

et al. 2019

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“…These ribozymes were originally selected to react with biotinyl-Tyr(Me)-oxazolone (BYO), a chemically activated amino acid. The 5(4H)-oxazolones and related Ncarboxyanhydrides can be made abiotically under prebiotically plausible conditions [40][41][42][43][44][45][46][47][48] . Three distinct, evolutionarily unrelated catalytic motifs had been discovered from the exhaustive search.…”

Section: Introductionmentioning

confidence: 99%

Error minimization and specificity could emerge in a genetic code as by-products of prebiotic evolution

Janzen

Shen

Liu

et al. 2021

Preprint

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The emergence of the genetic code was a major transition in the evolution from a prebiotic RNA world to the earliest modern cells. A prominent feature of the standard genetic code is error minimization, or the tendency of mutations to be unusually conservative in preserving biophysical features of the amino acid. While error minimization is often assumed to result from natural selection, it has also been speculated that error minimization may be a by-product of emergence of the genetic code. During establishment of the genetic code in an RNA world, self-aminoacylating ribozymes would enforce the mapping of amino acids to anticodons. Here we show that expansion of the genetic code, through co-option of ribozymes for new substrates, could result in error minimization as an emergent property. Using self-aminoacylating ribozymes previously identified during an exhaustive search of sequence space, we measured the activity of thousands of candidate ribozymes on alternative substrates (activated analogs for tryptophan, phenylalanine, leucine, isoleucine, valine, and methionine). Related ribozymes exhibited preferences for biophysically similar substrates, indicating that co-option of existing ribozymes to adopt additional amino acids into the genetic code would itself lead to error minimization. Furthermore, ribozyme activity was positively correlated with specificity, indicating that selection for increased activity would also lead to increased specificity. These results demonstrate that by-products of the evolution and functional expansion of a ribozyme system could lead to adaptive properties of a genetic code. Such 'spandrels' could thus underlie significant prebiotic developments.

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“…However, the latter reaction competes with the rapid conversion of the aminoacyl adenylate intermediate into an N-carboxyanhydride (NCA) in the presence of CO2 3,4 . NCAs are inefficient reagents for the direct aminoacylation of ribonucleotides 5 . High yielding aminoacylation pathways employing NCAs 6 or in situ activation chemistry 7 are known, but they require a 3′phosphate moiety, and are generally limited to N-blocked amino acids.…”

Section: Introductionmentioning

confidence: 99%

A Potential Role for Aminoacylation in Primordial RNA Copying Chemistry

Radakovic

Wright²,

Lelyveld³

et al. 2020

Preprint

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Aminoacylated tRNAs are the substrates for ribosomal protein synthesis in all branches of life, implying an ancient origin for aminoacylation chemistry. In the 1970s, Orgel and colleagues reported potentially prebiotic routes to aminoacylated nucleotides and their RNA templated condensation to form amino acid bridged dinucleotides. However, it is unclear whether such reactions would have aided or impeded nonenzymatic RNA replication. Determining whether aminoacylated RNAs could have been advantageous in evolution prior to the emergence of protein synthesis remains a key challenge. We therefore tested the ability of aminoacylated RNA to participate in both templated primer extension and ligation reactions. We find that at low magnesium concentrations that favor fatty acid-based protocells, these reactions proceed orders of magnitude more rapidly than when initiated from the cis-diol of unmodified RNA. We further demonstrate that amino acid bridged RNAs can act as templates in a subsequent round of copying. Our results suggest that aminoacylation facilitated nonenzymatic RNA replication, thus outlining a potentially primordial functional link between aminoacylation chemistry and RNA replication.Abstract Figure

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5(4H)-Oxazolones as Effective Aminoacylation Reagents for the 3′-Terminus of RNA

Cited by 8 publications

References 13 publications

Mapping a Systematic Ribozyme Fitness Landscape Reveals a Frustrated Evolutionary Network for Self-Aminoacylating RNA

Mapping a Systematic Ribozyme Fitness Landscape Reveals a Frustrated Evolutionary Network for Self-Aminoacylating RNA

Error minimization and specificity could emerge in a genetic code as by-products of prebiotic evolution

A Potential Role for Aminoacylation in Primordial RNA Copying Chemistry

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