Template-directed Chemical Ligation to Obtain 3′-3′ and 5′-5′ Phosphodiester DNA Linkages

Chen, Haodong; Du, Feng; Chen, Gangyi; Streckenbach, Frank; Yasmeen, Aneela; Zhao, Yun; Tang, Zhuo

doi:10.1038/srep04595

Cited by 7 publications

(9 citation statements)

References 28 publications

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“…Finally, we demonstrate that N -cyano-2-aminoimidazole ( 2NH 2 ImCN ) is capable of activating CMP to furnish (Scheme 2) cytidine-5′-phosphoro-(2-aminoimidazole) ( 2NH 2 ImpC ). Several reports in the literature suggest that N -cyanoimidazole can serve 22,23 as an activating agent for the formation of phosphodiester bonds. 2NH 2 ImCN appeared attractive as a potentially prebiotic activating agent, because it represents one of the simplest possible extensions of the chemistry reported here, i.e., oxidative coupling with hydrogen cyanide.…”

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confidence: 99%

Common and Potentially Prebiotic Origin for Precursors of Nucleotide Synthesis and Activation

et al. 2017

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We have recently shown that 2-aminoimidazole is a superior nucleotide activating group for nonenzymatic RNA copying. Here we describe a prebiotic synthesis of 2-aminoimidazole that shares a common mechanistic pathway with that of 2-aminooxazole, a previously described key intermediate in prebiotic nucleotide synthesis. In the presence of glycolaldehyde, cyanamide, phosphate and ammonium ion, both 2-aminoimidazole and 2-aminooxazole are produced, with higher concentrations of ammonium ion and acidic pH favoring the former. Given a 1:1 mixture of 2-aminoimidazole and 2-aminooxazole, glyceraldehyde preferentially reacts and cyclizes with the latter, forming a mixture of pentose aminooxazolines, and leaving free 2-aminoimidazole available for nucleotide activation. The common synthetic origin of 2-aminoimidazole and 2-aminooxazole and their distinct reactivities are suggestive of a reaction network that could lead to both the synthesis of RNA monomers and to their subsequent chemical activation.

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confidence: 99%

Common and Potentially Prebiotic Origin for Precursors of Nucleotide Synthesis and Activation

et al. 2017

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“…21,30 Although some groups have reported template-directed ligation of DNA with the CNIm/Ni 2+ system, efficiencies were not as high as we observed using Cd 2+ . 33,35,37 Since the 5′-phosphorylated DNA can be easily prepared by incubation with a kinase and ATP, this rapid chemical ligation system should be widely applicable.…”

Section: ■ Results and Discussionmentioning

confidence: 99%

“…Of note, both 3′-phosphorylated DNA and 5′-phosphorylated DNA were efficiently ligated: ligation was quantitative after a 4 h incubation with 5 mM Cd 2+ and after a 1 h incubation with 20 mM Cd 2+ at 25 °C (Figure S3). Thus, ligation in the presence of CNIm and Cd 2+ is much faster than EDC-mediated chemical ligation. , Although some groups have reported template-directed ligation of DNA with the CNIm/Ni 2+ system, efficiencies were not as high as we observed using Cd 2+ . ,, Since the 5′-phosphorylated DNA can be easily prepared by incubation with a kinase and ATP, this rapid chemical ligation system should be widely applicable.…”

Section: Resultsmentioning

confidence: 99%

“…A side reaction induced by EDC is also problematic . Cyanogen bromide (BrCN) has also been used to facilitate DNA ligation, but the high concentration required (100–500 mM), HBr released, and side reactions prevent practical use. ,, Kanaya and Yanagawa reported that N -cyanoimidazole (CNIm) generated by a BrCN/M 2+ /imidazole system enabled effective chemical ligation with a relatively low amount of reagent (∼20 mM). , The ligation reaction by CNIm proceeds in the presence of Mn 2+ or Zn 2+ . − Mariani and Sutherland performed RNA ligation in the CNIm/Mn 2+ system for the elucidation of only 3′,5′-linked RNA emergence from the heterogeneous mixture . However, the reaction is slow, and the mechanism remains unknown.…”

Section: Introductionmentioning

confidence: 99%

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Rapid Chemical Ligation of DNA and Acyclic Threoninol Nucleic Acid (aTNA) for Effective Nonenzymatic Primer Extension

et al. 2023

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Previously, nonenzymatic primer extension reaction of acyclic l-threoninol nucleic acid (L-aTNA) was achieved in the presence of N-cyanoimidazole (CNIm) and Mn2+; however, the reaction conditions were not optimized and a mechanistic insight was not sufficient. Herein, we report investigation of the kinetics and reaction mechanism of the chemical ligation of L-aTNA to L-aTNA and of DNA to DNA. We found that Cd2+, Ni2+, and Co2+ accelerated ligation of both L-aTNA and DNA and that the rate-determining step was activation of the phosphate group. The activation was enhanced by duplex formation between a phosphorylated L-aTNA fragment and template, resulting in unexpectedly more effective L-aTNA ligation than DNA ligation. Under optimized conditions, an 8-mer L-aTNA primer could be elongated by ligation to L-aTNA trimers to produce a 29-mer full-length oligomer with 60% yield within 2 h at 4 °C. This highly effective chemical ligation system will allow construction of artificial genomes, robust DNA nanostructures, and xeno nucleic acids for use in selection methods. Our findings also shed light on the possible pre-RNA world.

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“…A condensing reagent N -cyanoimidazole (CNIm) can connect a phosphate with a neighboring hydroxyl group to generate a phosphodiester bond, the same type of bond formed with natural enzymatic ligation 50 , 51 . For DNA, ligation of various types of structures were achieved using CNIm, due to relatively high reaction yields 52 – 56 . We therefore expected that CNIm could be used to ligate l - a TNA.…”

Section: Introductionmentioning

confidence: 99%

Nonenzymatic polymerase-like template-directed synthesis of acyclic l-threoninol nucleic acid

et al. 2021

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Evolution of xeno nucleic acid (XNA) world essentially requires template-directed synthesis of XNA polymers. In this study, we demonstrate template-directed synthesis of an acyclic XNA, acyclicl-threoninol nucleic acid (l-aTNA), via chemical ligation mediated by N-cyanoimidazole. The ligation of an l-aTNA fragment on an l-aTNA template is significantly faster and occurs in considerably higher yield than DNA ligation. Both l-aTNA ligation on a DNA template and DNA ligation on an l-aTNA template are also observed. High efficiency ligation of trimer l-aTNA fragments to a template-bound primer is achieved. Furthermore, a pseudo primer extension reaction is demonstrated using a pool of random l-aTNA trimers as substrates. To the best of our knowledge, this is the first example of polymerase-like primer extension of XNA with all four nucleobases, generating phosphodiester bonding without any special modification. This technique paves the way for a genetic system of the l-aTNA world.

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Template-directed Chemical Ligation to Obtain 3′-3′ and 5′-5′ Phosphodiester DNA Linkages

Cited by 7 publications

References 28 publications

Common and Potentially Prebiotic Origin for Precursors of Nucleotide Synthesis and Activation

Common and Potentially Prebiotic Origin for Precursors of Nucleotide Synthesis and Activation

Rapid Chemical Ligation of DNA and Acyclic Threoninol Nucleic Acid (aTNA) for Effective Nonenzymatic Primer Extension

Nonenzymatic polymerase-like template-directed synthesis of acyclic l-threoninol nucleic acid

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