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Cited by 63DNA and Nucleic Acid ChemistryRNA and protein synthesis mechanismsAdvanced biosensing and bioanalysis techniques

The effect of leaving groups on binding and reactivity in enzyme-free copying of DNA and RNA

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Kervio, Eric, Sosson, Marilyne, Richert, Clemens·2016·Nucleic Acids Research
DOI 10.1093/nar/gkw476PMC PMC4937335
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Abstract

The template-directed incorporation of nucleotides at the terminus of a growing primer is the basis of the transmission of genetic information. Nature uses polymerases-catalyzed reactions, but enzyme-free versions exist that employ nucleotides with organic leaving groups. The leaving group affects yields, but it was not clear whether inefficient extensions are due to poor binding, low reactivity toward the primer, or rapid hydrolysis. We have measured the binding of a total of 15 different activated nucleotides to DNA or RNA sequences. Further, we determined rate constants for the chemical step of primer extension involving methylimidazolides or oxyazabenzotriazolides of deoxynucleotides or ribonucleotides. Binding constants range from 10 to >500 mM and rate constants from 0.1 to 370 M(-1) h(-1) For aminoterminal primers, a fast covalent step and slow hydrolysis are the main factors leading to high yields. For monomers with weakly pairing bases, the leaving group can improve binding significantly. A detailed mechanistic picture emerges that explains why some enzyme-free primer extensions occur in high yield, while others remain recalcitrant to copying without enzymatic catalysis. A combination of tight binding and rapid extension, coupled with slow hydrolysis induces efficient enzyme-free copying.

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  • The Richert laboratory noted the formation of an imidazolium-bridged dinucleotide and its high reactivity in primer extension reactions.

    “The Richert laboratory also noted the formation of an imidazolium-bridged dinucleotide and its high reactivity in primer extension reactions”

    From Protocells and RNA Self-Replication (2018) · cites this paper as Kervio et al. 2016

    The Results (page unavailable) report a side product tentatively assigned to an imidazoliumbisphosphate linking two nucleotides. They also report an initial burst of primer extension when that side product was present, with slower kinetics after purification. This supports the reported reactivity, but the claim presents the dinucleotide’s identity as established rather than tentative. (LLM, conf=0.83)

    Experimental — accuracy not yet measured

  • 1-Hydroxy-7-azabenzotriazole (HOAt) functions as an anionic leaving group and results in rapid primer extension attributable to direct reaction of the primer with the HOAt-activated monomer.

    “1-Hydroxy-7-azabenzotriazole (HOAt) functions in this manner and results in rapid primer extension attributable to direct reaction of the primer with the HOAt-activated monomer”

    From Protocells and RNA Self-Replication (2018) · cites this paper as Kervio et al. 2016

    The Introduction identifies oxyazabenzotriazole (OAt) as a leaving group, and the Results report faster covalent primer extension with OAt-activated deoxynucleotides than with methylimidazolide analogues. The Discussion describes a bound monomer reacting with the primer, but our copy does not establish the claim’s specific characterization of HOAt as an anionic leaving group; it also attributes the increased rate and yield to both improved binding and faster extension. (LLM, conf=0.83)

    Experimental — accuracy not yet measured

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Research works printed but not linked: 28

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Authors
Kervio, E., Sosson, M., & Richert, C.
Affiliations
University of Stuttgart (DE)
Corresponding author
Richert, Clemens
ORCID
Venue
Nucleic Acids Research
Publisher
Oxford University Press (OUP)
Published
2016-05-27
Volume
44
Issue
12
Pages
5504-5514
ISSN
  • 0305-1048 (ISSN-L)
  • 1362-4962
DOI
10.1093/nar/gkw476
PMC
PMC4937335
OpenAlex
W2404300937
Cited by
63
References
10
Citations by year
2026: 2 · 2025: 4 · 2024: 5 · 2023: 4 · 2022: 6 · 2021: 7 · 2020: 9 · 2019: 5 · 2018: 7 · 2017: 9 · 2016: 4 · 2014: 1
FWCI
4.51
Type
journal article
Language
en
Open access
Licence
Locations
  • Nucleic Acids Research (publishedVersion, cc-by-nc)
  • PubMed Central (publishedVersion, cc-by-nc)
  • PubMed (publishedVersion)
  • HighWire Press Open Archive (submittedVersion)
Funders
Deutsche Forschungsgemeinschaft (RI 1063/8-2)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › DNA and Nucleic Acid Chemistry
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA and protein synthesis mechanisms
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Advanced biosensing and bioanalysis techniques
Keywords
leaving groups, primer extension, nucleotide hydrolysis
MeSH terms
Base Sequence, DNA, DNA Replication, Hydrolysis, Kinetics, Ribonucleotides, RNA, DNA Primers

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