Limits of RNA 2′-OH Mimicry by Fluorine: Crystal Structure of <i>Bacillus halodurans</i> RNase H Bound to a 2′-FRNA:DNA Hybrid

Pallan, Pradeep S.; Prakash, Thazha P.; Leon, Arnie R. de; Egli, Martin

doi:10.1021/acs.biochem.6b00849

Cited by 13 publications

(12 citation statements)

References 26 publications

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“…Previous examples have shown that the presence of small modifications can result in significant changes in reactivity with some reports of note. (1) The loss of activity of RNase H has been reported upon substitution of RNA with 2′-fluoro substituents (known to exist in the 3′-endo conformation), attributed to electronic and geometric differences, 67 and (2) it has also been shown that cleavage is shifted if the DNA strand contains oxidatively induced modifications 68 or that the presence of ethyl groups along the backbone of the 20-mer that is used shifts the degradation patterns away from these sites. 69 Importantly, these examples also used RNase H from E. coli and point to the importance of the structural aspects within the RNA:DNA duplexes to maintain fidelity, efficiency, and selectivity of this ubiquitous ribonuclease.…”

Section: ■ Discussionmentioning

confidence: 99%

Reactivity and Specificity of RNase T₁, RNase A, and RNase H toward Oligonucleotides of RNA Containing 8-Oxo-7,8-dihydroguanosine

et al. 2018

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Understanding how oxidatively damaged RNA interacts with ribonucleases is important because of its proposed role in the development and progression of disease. Thus, understanding structural aspects of RNA containing lesions generated under oxidative stress, as well as its interactions with other biopolymers, is fundamental. We explored the reactivity of RNase A, RNase T, and RNase H toward oligonucleotides of RNA containing 8-oxo-7,8-dihydroguanosine (8oxoG). This is the first example that addresses this relationship and will be useful for understanding (1) how these RNases can be used to characterize the structural impact that this lesion has on RNA and (2) how oxidatively modified RNA may be handled intracellularly. 8-OxoG was incorporated into 10-16-mers of RNA, and its reactivity with each ribonuclease was assessed via electrophoretic analyses, circular dichroism, and the use of other C8-purine-modified analogues (8-bromoguanosine, 8-methoxyguanosine, and 8-oxoadenosine). RNase T does not recognize sites containing 8-oxoG, while RNase A recognizes and cleaves RNA at positions containing this lesion while differentiating if it is involved in H-bonding. The selectivity of RNase A followed the order C > 8-oxoG ≈ U. In addition, isothermal titration calorimetry showed that an 8-oxoG-C3'-methylphosphate derivative can inhibit RNase A activity. Cleavage patterns obtained from RNase H displayed changes in reactivity in a sequence- and concentration-dependent manner and displayed recognition at sites containing the modification in some cases. These data will aid in understanding how this modification affects reactivity with ribonucleases and will enable the characterization of global and local structural changes in oxidatively damaged RNA.

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Section: ■ Discussionmentioning

confidence: 99%

Reactivity and Specificity of RNase T₁, RNase A, and RNase H toward Oligonucleotides of RNA Containing 8-Oxo-7,8-dihydroguanosine

et al. 2018

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“…However, there are limits to the bioisosteric relationship between a 2′-fluoro and a 2′-hydroxyl substituent in the context of RNA. While the use of fluorinated RNA is common in siRNA and aptamers as a means of stabilizing the RNA, incorporation into RNA:DNA hybrids has been found to be poorly efficacious toward activating mammalian RNaseH …”

Section: C–f As a Carbinol (C–oh) Mimicmentioning

confidence: 99%

“…While the use of fluorinated RNA is common in siRNA and aptamers as a means of stabilizing the RNA, incorporation into RNA:DNA hybrids has been found to be poorly efficacious toward activating mammalian RNaseH. 191 Collagen is the most abundant protein in animals, amounting to ∼30% of the total protein content of humans and ∼75% of the weight of human skin. The polypeptide chains of collagen are comprised of approximately 300 repeats of the triplet sequence XYG in which X is often an L-Pro residue while Y is frequently a 4-(R)-hydroxy-L-proline (438, Hyp) residue.…”

Section: Fluorinated Amines and Mimesis Of Imide-like Motifsmentioning

confidence: 99%

Fluorine and Fluorinated Motifs in the Design and Application of Bioisosteres for Drug Design

2018

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The electronic properties and relatively small size of fluorine endow it with considerable versatility as a bioisostere and it has found application as a substitute for lone pairs of electrons, the hydrogen atom, and the methyl group while also acting as a functional mimetic of the carbonyl, carbinol, and nitrile moieties. In this context, fluorine substitution can influence the potency, conformation, metabolism, membrane permeability, and P-gp recognition of a molecule and temper inhibition of the hERG channel by basic amines. However, as a consequence of the unique properties of fluorine, it features prominently in the design of higher order structural metaphors that are more esoteric in their conception and which reflect a more sophisticated molecular construction that broadens biological mimesis. In this Perspective, applications of fluorine in the construction of bioisosteric elements designed to enhance the in vitro and in vivo properties of a molecule are summarized.

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“…For example, a 2′-F containing RNA can be prepared by enzymatic transcription using the same DNA template but with 2′-F nucleotides. However, a chemically modified RNA with the sequence identical to its unmodified RNA aptamer could have a very different structure and consequently a much attenuated potency, as compared with the canonical RNA. − The second approach is generally more desirable in which SELEX is used with a chemically modified library to isolate a chemically modified aptamer: the chemical modifications are introduced to the library or “pre-SELEX”. By this approach, a chemically modified RNA aptamer with the best fitted structure could be selected from a large RNA library (e.g., ∼10 14 sequences) without compromising the potency …”

Section: Introductionmentioning

confidence: 99%

Chemically Modified, α-Amino-3-hydroxy-5-methyl-4-isoxazole (AMPA) Receptor RNA Aptamers Designed for in Vivo Use

Huang

Wen

et al. 2017

ACS Chem. Neurosci.

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Glutamate ion channels have three subtypes, that is, α-amino-3-hydroxy-5-methyl-4-isoxazole (AMPA), kainate, and N-methyl-d-aspartate (NMDA) receptors. Excessive activity of these receptor subtypes either individually or collectively is involved in various neurological disorders. RNA aptamers as antagonists of these receptors are potential therapeutics. For developing aptamer therapeutics, the RNA aptamers must be chemically modified to become ribonuclease-resistant or stable in biological fluids. Using systematic evolution of ligands by exponential enrichment (SELEX) and a chemically modified library, prepared enzymatically (i.e., the library contains RNAs with 2'-fluoro modified nucleoside triphosphates or ATPs, CTPs and UTPs, but regular GTPs), we have isolated an aptamer. The short aptamer (69 nucleotides) FN1040s selectively inhibits the GluA1 and GluA2Q AMPA receptor subunits, whereas the full-length aptamer (101 nucleotides) FN1040 additionally inhibits GluK1, but not GluK2, kainate receptor, and GluN1a/2A and GluN1a/2B, the two major native NMDA receptors. The two aptamers show similar potency (2-4 μM) and are stable with a half-life of at least 2 days in serum-containing medium or cerebrospinal fluid. Therefore, these two aptamers are amenable for in vivo use.

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Limits of RNA 2′-OH Mimicry by Fluorine: Crystal Structure of Bacillus halodurans RNase H Bound to a 2′-FRNA:DNA Hybrid

Cited by 13 publications

References 26 publications

Reactivity and Specificity of RNase T₁, RNase A, and RNase H toward Oligonucleotides of RNA Containing 8-Oxo-7,8-dihydroguanosine

Reactivity and Specificity of RNase T₁, RNase A, and RNase H toward Oligonucleotides of RNA Containing 8-Oxo-7,8-dihydroguanosine

Fluorine and Fluorinated Motifs in the Design and Application of Bioisosteres for Drug Design

Chemically Modified, α-Amino-3-hydroxy-5-methyl-4-isoxazole (AMPA) Receptor RNA Aptamers Designed for in Vivo Use

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Limits of RNA 2′-OH Mimicry by Fluorine: Crystal Structure of Bacillus halodurans RNase H Bound to a 2′-FRNA:DNA Hybrid

Cited by 13 publications

References 26 publications

Reactivity and Specificity of RNase T1, RNase A, and RNase H toward Oligonucleotides of RNA Containing 8-Oxo-7,8-dihydroguanosine

Reactivity and Specificity of RNase T1, RNase A, and RNase H toward Oligonucleotides of RNA Containing 8-Oxo-7,8-dihydroguanosine

Fluorine and Fluorinated Motifs in the Design and Application of Bioisosteres for Drug Design

Chemically Modified, α-Amino-3-hydroxy-5-methyl-4-isoxazole (AMPA) Receptor RNA Aptamers Designed for in Vivo Use

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Product

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Reactivity and Specificity of RNase T₁, RNase A, and RNase H toward Oligonucleotides of RNA Containing 8-Oxo-7,8-dihydroguanosine

Reactivity and Specificity of RNase T₁, RNase A, and RNase H toward Oligonucleotides of RNA Containing 8-Oxo-7,8-dihydroguanosine