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Volume 193, 2016
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Energetics of base flipping at a DNA mismatch site confined at the latch constriction of α-hemolysin

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Abstract

Unique, two-state modulating current signatures are observed when a cytosine–cytosine mismatch pair is confined at the 2.4 nm latch constriction of the α-hemolysin (αHL) nanopore. We have previously speculated that the modulation is due to base flipping at the mismatch site. Base flipping is a biologically significant mechanism in which a single base is rotated out of the DNA helical stack by 180°. It is the mechanism by which enzymes are able to access bases for repair operations without disturbing the global structure of the helix. Here, temperature dependent ion channel recordings of individual double-stranded DNA duplexes inside αHL are used to derive thermodynamic (ΔH, ΔS) and kinetic (EA) parameters for base flipping of a cytosine at an unstable cytosine–cytosine mismatch site. The measured activation energy for flipping a cytosine located at the latch of αHL out of the helix (18 ± 1 kcal mol−1) is comparable to that previously reported for base flipping at mismatch sites from NMR measurements and potential mean force calculations. We propose that the αHL nanopore is a useful tool for measuring conformational changes in dsDNA at the single molecule level.

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Publication details

The article was received on 21 Mar 2016, accepted on 29 Mar 2016 and first published on 29 Mar 2016


Article type: Paper
DOI: 10.1039/C6FD00058D
Citation: Faraday Discuss., 2016,193, 471-485
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    Energetics of base flipping at a DNA mismatch site confined at the latch constriction of α-hemolysin

    R. P. Johnson, R. T. Perera, A. M. Fleming, C. J. Burrows and H. S. White, Faraday Discuss., 2016, 193, 471
    DOI: 10.1039/C6FD00058D

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