Structural basis for the unusual properties of 2′,5′ nucleic acids and their complexes with RNA and DNA

Premraj, Bernard J.; Raja, Swaminathan; Yathindra, N.

doi:10.1016/s0301-4622(02)00040-6

Cited by 14 publications

(18 citation statements)

References 68 publications

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“…However, the mechanism responsible for this destabilization has not yet been satisfactorily elucidated, although a very preliminary modeling study has suggested that the reduced base overlap between adjacent intrastrand bases caused by the 2′-5′ linkage might be one of the reasons for the decreased T m (17). In addition, the diminished chemical stability of this linkage in the duplex state has been suggested as a potential proofreading mechanism for linkage heterogeneity in prebiotic RNA synthesis (5).…”

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confidence: 99%

Structural insights into the effects of 2′-5′ linkages on the RNA duplex

Sheng

Engelhart

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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The mixture of 2′-5′ and 3′-5′ linkages generated during the nonenzymatic replication of RNA has long been regarded as a central problem for the origin of the RNA world. However, we recently observed that both a ribozyme and an RNA aptamer retain considerable functionality in the presence of prebiotically plausible levels of linkage heterogeneity. To better understand the RNA structure and function in the presence of backbone linkage heterogeneity, we obtained high-resolution X-ray crystal structures of a native 10-mer RNA duplex (1.32 Å) and two variants: one containing one 2′-5′ linkage per strand (1.55 Å) and one containing three such linkages per strand (1.20 Å). We found that RNA duplexes adjust their local structures to accommodate the perturbation caused by 2′-5′ linkages, with the flanking nucleotides buffering the disruptive effects of the isomeric linkage and resulting in a minimally altered global structure. Although most 2′-linked sugars were in the expected 2′-endo conformation, some were partially or fully in the 3′-endo conformation, suggesting that the energy difference between these conformations was relatively small. Our structural and molecular dynamic studies also provide insight into the diminished thermal and chemical stability of the duplex state associated with the presence of 2′-5′ linkages. Our results contribute to the view that a low level of 2′-5′ substitution would not have been fatal in an early RNA world and may in contrast have been helpful for both the emergence of nonenzymatic RNA replication and the early evolution of functional RNAs.origin of life | backbone heterogeneity | X-ray crystallography T he capacity of RNA to act as both a carrier of genetic information and as a catalyst has led many to investigate its potential role as the first biopolymer (1-4). An early stage involving nonenzymatic replication simplifies RNA-first scenarios, but known nonenzymatic copying reactions generate a mixture of 3′-5′ and 2′-5′ backbone linkages because of the similar nucleophilicity and orientation of the 2′ and 3′ hydroxyl groups on ribose (Fig. 1). Although regioselectivity for the 3′-5′ linkage can be improved by using different metal ions or activated monomers, it reaches, at most, ∼90% (5-11). This lack of regiospecificity has been regarded as a central problem for the emergence of the RNA world, because the resulting backbone heterogeneity was expected to disrupt the folding, molecular recognition, and catalytic properties of functional RNAs. However, we recently observed that functional nucleic acid molecules can still evolve in the presence of nonheritable mixed DNA/RNA backbone heterogeneity (12), and known functional RNAs retain catalytic and ligand binding behavior in the presence of 2′-5′/3′-5′ backbone linkage heterogeneity (13).The well-known duplex-destabilizing property of 2′-5′ linkages can enable thermal strand separation of long RNA duplexes in the presence of the high Mg 2+ concentrations required for known prebiotic copying reactions (13-16). However, the mechanism r...

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confidence: 99%

Structural insights into the effects of 2′-5′ linkages on the RNA duplex

Sheng

Engelhart

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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“…In contrast, we find that structural mutagenesis of G14, whereby guanine functional groups are retained but a geometric perturbation of the active site is introduced,20 results in a comparatively mild reduction in 10–23 activity (around fivefold; Figure 4 b,c). …”

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confidence: 74%

Enzymatic Synthesis of Nucleic Acids with Defined Regioisomeric 2′‐5′ Linkages

et al. 2015

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Information‐bearing nucleic acids display universal 3′‐5′ linkages, but regioisomeric 2′‐5′ linkages occur sporadically in non‐enzymatic RNA synthesis and may have aided prebiotic RNA replication. Herein we report on the enzymatic synthesis of both DNA and RNA with site‐specific 2′‐5′ linkages by an engineered polymerase using 3′‐deoxy‐ or 3′‐O‐methyl‐NTPs as substrates. We also report the reverse transcription of the resulting modified nucleic acids back to 3′‐5′ linked DNA with good fidelity. This enables a fast and simple method for “structural mutagenesis” by the position‐selective incorporation of 2′‐5′ linkages, whereby nucleic acid structure and function may be probed through local distortion by regioisomeric linkages while maintaining the wild‐type base sequence as we demonstrate for the 10–23 RNA endonuclease DNAzyme.

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“…Diese "Primerabtastung" ermçglicht eine schnelle Bestimmung von obligatorischen 3'-5'-Verknüpfungen, die empfindlich auf strukturelle Verformungen reagieren, neutralen Positionen, die sowohl eine 3'-5'-a ls auch eine 2'-5'-Geometrie tolerieren, und allen Positionen, für die das Einfügen von 2'-5'-Verknüpfungen vorteilhaft ist. [19] Dagegen stellten wir fest, dass eine strukturelle Mutagenese von G14 -w obei Guanin als funktionelle Gruppe erhalten bleibt, aber eine geometrische Stçrung in das aktive Zentrum einfügt wird [20] -e ine vergleichsweise milde Verringerung der 10-23-Aktivität (ca. Beispielsweise zeigte die Mutagenese des distalen Substrat-Bindearms, dass das Einfügen von 2'-5'-Verknüpfungen nach den Nukleotiden G18 und G22 zu einer moderaten Aktivitätszunahme führt (Abbildung 4c), vermutlich aufgrund einer reduzierten Produktinhibierung.…”

Section: Methodsunclassified

Enzymatische Synthese von Nukleinsäuren mit definierten regioisomeren 2′‐5′‐Verknüpfungen

et al. 2015

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Informationentragende Nukleinsäuren sind universell über 3'-5'-Bindungen verknüpft. Regioisomere 2'-5'-Verknüpfungen bilden sich dagegen unregelmäßig während der nichtenzymatischen RNA-Synthese und waren womçglich hilfreich für die präbiotischeRNA-Replikation. Hier berichten wir über die enzymatischeS ynthese von DNAu nd RNAm it ortsspezifischen 2'-5'-Verknüpfungen durch ein gezielt verändertes Polymerase-Enzym, das 3'-Desoxy-oder 3'-O-Methyl-NTPs als Substrat verwendet. Zusätzlich vermelden wir die reverse Transkription der so hergestellten, modifizierten Nukleinsäuren zurück in 3'-5'-verknüpfte DNAm it guter Fidelität. Somit ergibt sich eine schnelle und einfache Methode zur "strukturellen Mutagenese" durch positionsselektives Einfügen von 2'-5'-Verknüpfungen, durch die,o hne ¾nderung der Basensequenz, die Struktur und Funktion von Nukleinsäuren anhand lokaler Deformationen durch regioisomere Bindungen untersucht werden kann. Beispielhaft wenden wir diese Methode auf das 10-23-RNA-Endonuklease-DNAzym an.

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Structural basis for the unusual properties of 2′,5′ nucleic acids and their complexes with RNA and DNA

Cited by 14 publications

References 68 publications

Structural insights into the effects of 2′-5′ linkages on the RNA duplex

Structural insights into the effects of 2′-5′ linkages on the RNA duplex

Enzymatic Synthesis of Nucleic Acids with Defined Regioisomeric 2′‐5′ Linkages

Enzymatische Synthese von Nukleinsäuren mit definierten regioisomeren 2′‐5′‐Verknüpfungen

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