Determination of the secondary structure of group II bulge loops using the fluorescent probe 2-aminopurine

Dishler, Abigael L.; McMichael, Elizabeth L.; Serra, Martin J.

doi:10.1261/rna.048306.114

Cited by 3 publications

(3 citation statements)

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“…2-Aminopurine (2AP) is a fluorescent analogue of adenine and is an excellent probe of local structural dynamics because 2AP can form a Watson–Crick-like 2AP-U pair and the 2AP fluorescence intensity is sensitive to its local structural environment, with single-stranded 2AP residues showing enhanced fluorescence compared to that of base-paired ones. − A mainly stacked-in bulge A with the bottom stem containing only one single G-C pair was observed in NMR studies for WT-25nt . We substituted the adenine at position −3 (A–3) in WT-25nt, +19G-25nt, WT-30nt, and +19G-30nt with 2AP (Figure B) to further confirm that the +19G mutation induces the structural rearrangement in the +19G-30nt hairpin.…”

Section: Resultsmentioning

confidence: 99%

A Disease-Causing Intronic Point Mutation C19G Alters Tau Exon 10 Splicing via RNA Secondary Structure Rearrangement

et al. 2019

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Alternative splicing of MAPT cassette exon 10 produces tau isoforms with four microtubule-binding repeat domains (4R) upon exon inclusion or three repeats (3R) upon exon skipping. In human neurons, deviations from the ∼1:1 physiological 4R:3R ratio lead to frontotemporal dementia with Parkinsonism linked to chromosome 17 (FTDP-17). Certain FTDP-17-associated mutations affect a regulatory hairpin that sequesters the exon 10 5′ splice site (5′ss, located at the exon 10–intron 10 junction). These mutations tend to increase the 4R:3R ratio by destabilizing the hairpin, thereby improving 5′ss recognition by U1 snRNP. Interestingly, a single C-to-G mutation at the 19th nucleotide in intron 10 (C19G or +19G) decreases the level of exon 10 inclusion significantly from 56% to 1%, despite the disruption of a G-C base pair in the bottom stem of the hairpin. Here, we show by biophysical characterization, including thermal melting, fluorescence, and single-molecule mechanical unfolding using optical tweezers, that the +19G mutation alters the structure of the bottom stem, resulting in the formation of a new bottom stem with enhanced stability. The cell culture alternative splicing patterns of a series of minigenes reveal that the splicing activities of the mutants with destabilizing mutations on the top stem can be compensated in a position-dependent manner by the +19G mutation in the bottom stem. We observed an excellent correlation between the level of exon 10 inclusion and the rate of mechanical unfolding at 10 pN, indicating that the unfolding of the splice site hairpins (to facilitate subsequent binding of U1 snRNA) may be aided by helicases or other proteins.

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Section: Resultsmentioning

confidence: 99%

A Disease-Causing Intronic Point Mutation C19G Alters Tau Exon 10 Splicing via RNA Secondary Structure Rearrangement

et al. 2019

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“…Here, the number of hydrogen bonds has not changed but the type of hydrogen bonds has changed. An RNA hairpin where an A-U to 2-aminopurine•U pair replacement occurred at the stem region had a difference in free energy of 0.8 kcal/mol ( 8 ). The range of values found in Tables 2 and 4 suggests that a single value representing the energy of a hydrogen bond may be insufficient for understanding the contribution to duplex stability by functional groups that participate in hydrogen bonds.…”

Section: Discussionmentioning

confidence: 99%

The loss of a hydrogen bond: Thermodynamic contributions of a non-standard nucleotide

Jolley

Znosko

2016

Nucleic Acids Res

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Non-standard nucleotides are ubiquitous in RNA. Thermodynamic studies with RNA duplexes containing non-standard nucleotides, whether incorporated naturally or chemically, can provide insight into the stability of Watson–Crick pairs and the role of specific functional groups in stabilizing a Watson–Crick pair. For example, an A-U, inosine•U and pseudouridine•A pair each form two hydrogen bonds. However, an RNA duplex containing a central I•U pair or central Ψ•A pair is 2.4 kcal/mol less stable or 1.7 kcal/mol more stable, respectively, than the corresponding duplex containing an A-U pair. In the non-standard nucleotide purine, hydrogen replaces the exocyclic amino group of A. This replacement results in a P•U pair containing only one hydrogen bond. Optical melting studies were performed with RNA duplexes containing P•U pairs adjacent to different nearest neighbors. The resulting thermodynamic parameters were compared to RNA duplexes containing A-U pairs in order to determine the contribution of the hydrogen bond involving the exocyclic amino group. Results indicate a loss of 1.78 kcal/mol, on average, when an internal P•U replaces A-U in an RNA duplex. This value is compared to the thermodynamics of a hydrogen bond determined by similar methods. Nearest neighbor parameters were derived for use in free energy and secondary structure prediction software.

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“…27 This follows similar conclusions for DNA bulges. 20,28 Melting temperatures for single bulges in RNA hairpins [29][30][31] as well as for longer bulges 32 further highlight these shortcommings. Therefore, there is a motivation to go beyond the nearestneighbor model but which are still computationally efficient to handle a large number of sequences.…”

Section: Introductionmentioning

confidence: 98%

An asymmetric mesoscopic model for single bulges in RNA

Martins

Weber

2017

The Journal of Chemical Physics

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Simple one-dimensional DNA or RNA mesoscopic models are of interest for their computational efficiency while retaining the key elements of the molecular interactions. However, they only deal with perfectly formed DNA or RNA double helices and consider the intra-strand interactions to be the same on both strands. This makes it difficult to describe highly asymmetric structures such as bulges and loops and, for instance, prevents the application of mesoscopic models to determine RNA secondary structures. Here we derived the conditions for the Peyrard-Bishop mesoscopic model to overcome these limitations and applied it to the calculation of single bulges, the smallest and simplest of these asymmetric structures. We found that these theoretical conditions can indeed be applied to any situation where stacking asymmetry needs to be considered. The full set of parameters for group I RNA bulges was determined from experimental melting temperatures using an optimization procedure, and we also calculated average opening profiles for several RNA sequences. We found that guanosine bulges show the strongest perturbation on their neighboring base pairs, considerably reducing the on-site interactions of their neighboring base pairs.

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Determination of the secondary structure of group II bulge loops using the fluorescent probe 2-aminopurine

Cited by 3 publications

References 62 publications

A Disease-Causing Intronic Point Mutation C19G Alters Tau Exon 10 Splicing via RNA Secondary Structure Rearrangement

A Disease-Causing Intronic Point Mutation C19G Alters Tau Exon 10 Splicing via RNA Secondary Structure Rearrangement

The loss of a hydrogen bond: Thermodynamic contributions of a non-standard nucleotide

An asymmetric mesoscopic model for single bulges in RNA

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