Artificial nucleotide codons for enzymatic DNA synthesis

Sabat, Nazarii; Stämpfli, Andreas; Flamme, Marie; Hanlon, Steven; Bisagni, Serena; Sladojevich, Filippo; Püntener, Kurt; Hollenstein, Marcel

doi:10.1039/d3cc04933g

Cited by 7 publications

(1 citation statement)

References 47 publications

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“…7 ). The replacement of internal phosphodiester bonds of trinucleotide triphosphates with phosphorothioates moieties permits to resist against the phosphatase activity of DNA polymerases and concomitantly supports enzymatic synthesis of DNA three-by-three 118 . While this method has only been evaluated on rather simple nucleotidic scaffolds so far, it could be extended to XNA-containing trinucleotides or codons capped with transient blocking groups at the 3’-termini for controlled enzymatic XNA synthesis.…”

Section: Mainmentioning

confidence: 99%

Controlled enzymatic synthesis of oligonucleotides

Pichon,

Hollenstein

2024

Commun Chem

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Oligonucleotides are advancing as essential materials for the development of new therapeutics, artificial genes, or in storage of information applications. Hitherto, our capacity to write (i.e., synthesize) oligonucleotides is not as efficient as that to read (i.e., sequencing) DNA/RNA. Alternative, biocatalytic methods for the de novo synthesis of natural or modified oligonucleotides are in dire need to circumvent the limitations of traditional synthetic approaches. This Perspective article summarizes recent progress made in controlled enzymatic synthesis, where temporary blocked nucleotides are incorporated into immobilized primers by polymerases. While robust protocols have been established for DNA, RNA or XNA synthesis is more challenging. Nevertheless, using a suitable combination of protected nucleotides and polymerase has shown promises to produce RNA oligonucleotides even though the production of long DNA/RNA/XNA sequences (>1000 nt) remains challenging. We surmise that merging ligase- and polymerase-based synthesis would help to circumvent the current shortcomings of controlled enzymatic synthesis.

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Section: Mainmentioning

confidence: 99%

Controlled enzymatic synthesis of oligonucleotides

Pichon,

Hollenstein

2024

Commun Chem

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Xeno Nucleic Acids as Functional Materials: From Biophysical Properties to Application

Bian,

Pei,

Gao

et al. 2024

Adv Healthcare Materials

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Xeno nucleic acid (XNA) are artificial nucleic acids, in which the chemical composition of the sugar moiety is changed. These modifications impart distinct physical and chemical properties to XNAs, leading to changes in their biological, chemical, and physical stability. Additionally, these alterations influence the binding dynamics of XNAs to their target molecules. Consequently, XNAs find expanded applications as functional materials in diverse fields. This review provides a comprehensive summary of the distinctive biophysical properties exhibited by various modified XNAs and explores their applications as innovative functional materials in expanded fields.

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2’,3’‐Protected Nucleotides as Building Blocks for Enzymatic de novo RNA Synthesis

Pichon,

Levi‐Acobas,

Kitoun

et al. 2024

Chemistry A European J

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Besides being a key player in numerous, fundamental biological process, RNA also represents a versatile platform for the creation of therapeutic agents and efficient vaccines. The production of RNA oligonucleotides, especially those decorated with chemical modifications, cannot meet the exponential demand. Due to the inherent limits of solid‐phase synthesis and in vitro transcription, alternative, biocatalytic approaches are in dire need to facilitate the production of RNA oligonucleotides. Here, we present a first step towards the controlled enzymatic synthesis of RNA oligonucleotides. We have explored the possibility of a simple protection step of the vicinal cis‐diol moiety to temporarily block ribonucleotides. We demonstrate that pyrimidine nucleotides protected with acetals, particularly 2',3'‐O‐isopropylidene, are well‐tolerated by the template‐independent RNA polymerase PUP (polyU polymerase) and highly efficient coupling reactions can be achieved within minutes – an important feature for the development of enzymatic de novo synthesis protocols. Even though purines are not equally well‐tolerated, these findings clearly demonstrate the possibility of using cis‐diol‐protected ribonucleotides combined with template‐independent polymerases for the stepwise construction of RNA oligonucleotides.

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Artificial nucleotide codons for enzymatic DNA synthesis

Abstract: Synthesis three-by-three: trinucleotide triphosphates can be used for enzymatic DNA synthesis provided their backbone is stabilized with chemical modifications.

Cited by 7 publications

References 47 publications

Controlled enzymatic synthesis of oligonucleotides

Controlled enzymatic synthesis of oligonucleotides

Xeno Nucleic Acids as Functional Materials: From Biophysical Properties to Application

2’,3’‐Protected Nucleotides as Building Blocks for Enzymatic de novo RNA Synthesis

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