Determination of the secondary structure of group II bulge loops using the fluorescent probe 2-aminopurine
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Issue Date
2015-03-24
Authors
Dishler, Abigael L.
McMichael, Elizabeth L.
Serra, Martin J.
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Keywords
bulge loops , secondary structure , thermodynamics
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Abstract
Eleven RNA hairpins containing 2-aminopurine (2-AP) in either base-paired or single nucleotide bulge loop positions were optically melted in 1 M NaCl; and, the thermodynamic parameters ΔH°, ΔS°, ΔG°37, and TM for each hairpin were determined. Substitution of 2-AP for an A (adenosine) at a bulge position (where either the 2-AP or A is the bulge) in the stem of a hairpin, does not affect the stability of the hairpin. For group II bulge loops such as AA/U, where there is ambiguity as to which of the A residues is paired with the U, hairpins with 2-AP substituted for either the 5′ or 3′ position in the hairpin stem have similar stability. Fluorescent melts were performed to monitor the environment of the 2-AP. When the 2-AP was located distal to the hairpin loop on either the 5′ or 3′ side of the hairpin stem, the change in fluorescent intensity upon heating was indicative of an unpaired nucleotide. A database of phylogenetically determined RNA secondary structures was examined to explore the presence of naturally occurring bulge loops embedded within a hairpin stem. The distribution of bulge loops is discussed and related to the stability of hairpin structures.
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Biochemistry
Chemistry
Chemistry
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© 2015 Dishler et al. This article, published in RNA, is available under a Creative Commons License (Attribution 4.0 International), as described at http://creativecommons.org/licenses/by/4.0/.
Citation
Dishler, Abigael L., Elizabeth L. McMichael, and Martin J. Serra. 2015. "Determination of the secondary structure of group II bulge loops using the fluorescent probe 2-aminopurine." RNA 21, no. 5: 975-984.
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Published article
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Cold Spring Harbor Laboratory Press