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Cleavage of fragments containing DNA mismatches by enzymic and chemical probes

Cleavage of fragments containing DNA mismatches by enzymic and chemical probes
Cleavage of fragments containing DNA mismatches by enzymic and chemical probes
We prepared synthetic 50-mer DNA duplexes, each containing four mismatched base-pairs in similar positions. We examined their cleavage by DNases I and II, micrococcal nuclease (MNase), methidiumpropyl-EDTA-Fe(II) [MPE-Fe(II)] and hydroxyl radicals. We find that single mismatches only produce subtle changes in the DNase I-cleavage pattern, the most common of which is attenuated cleavage at locations 2–3 bases on the 3´-side of the mismatch. Subtle changes are also observed in most of the DNase II-cleavage patterns, although GT and GG inhibit the cleavage over longer regions and generate patterns that resemble footprints. MNase cleaves the heteroduplexes at the mismatches themselves (except for CC), and in some cases cleaves CpG and CpC steps. None of the mismatches causes any change in the cleavage patterns produced by hydroxyl radicals or MPE-Fe(II). We also examined the cleavage patterns of fragments containing tandem GA mismatches in the sequences RGAY/RGAY and YGAR/YGAR (R, purine; Y, pyrimidine). RGAY causes only subtle changes in the cleavage patterns, which are similar to those seen with single mismatches, except that there are no changes in MNase cleavage. However, YGAR inhibits DNases I and II cleavage over 4–6 bases, and attenuates MPE-Fe(II) and hydroxyl radical cleavage at 2 bases. These changes suggest that this mismatch has a more pronounced effect on the local DNA structure. These changes are discussed in terms of the structural and dynamic effects of each mismatch.
DNA mismatch, DNase I, hydroxyl radical, methidiumpropyl-EDTA-Fe(II)
1470-8728
697-708
Brown, J.
64c8be68-e3cb-49ff-b5b0-525db5f4bcd0
Brown, T.
a64aae36-bb30-42df-88a2-11be394e8c89
Fox, K.R.
9da5debc-4e45-473e-ab8c-550d1104659f
Brown, J.
64c8be68-e3cb-49ff-b5b0-525db5f4bcd0
Brown, T.
a64aae36-bb30-42df-88a2-11be394e8c89
Fox, K.R.
9da5debc-4e45-473e-ab8c-550d1104659f

Brown, J., Brown, T. and Fox, K.R. (2003) Cleavage of fragments containing DNA mismatches by enzymic and chemical probes. Biochemical Journal, 371 (3), 697-708. (doi:10.1042/BJ20021847).

Record type: Article

Abstract

We prepared synthetic 50-mer DNA duplexes, each containing four mismatched base-pairs in similar positions. We examined their cleavage by DNases I and II, micrococcal nuclease (MNase), methidiumpropyl-EDTA-Fe(II) [MPE-Fe(II)] and hydroxyl radicals. We find that single mismatches only produce subtle changes in the DNase I-cleavage pattern, the most common of which is attenuated cleavage at locations 2–3 bases on the 3´-side of the mismatch. Subtle changes are also observed in most of the DNase II-cleavage patterns, although GT and GG inhibit the cleavage over longer regions and generate patterns that resemble footprints. MNase cleaves the heteroduplexes at the mismatches themselves (except for CC), and in some cases cleaves CpG and CpC steps. None of the mismatches causes any change in the cleavage patterns produced by hydroxyl radicals or MPE-Fe(II). We also examined the cleavage patterns of fragments containing tandem GA mismatches in the sequences RGAY/RGAY and YGAR/YGAR (R, purine; Y, pyrimidine). RGAY causes only subtle changes in the cleavage patterns, which are similar to those seen with single mismatches, except that there are no changes in MNase cleavage. However, YGAR inhibits DNases I and II cleavage over 4–6 bases, and attenuates MPE-Fe(II) and hydroxyl radical cleavage at 2 bases. These changes suggest that this mismatch has a more pronounced effect on the local DNA structure. These changes are discussed in terms of the structural and dynamic effects of each mismatch.

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More information

Submitted date: 27 November 2002
Published date: 1 May 2003
Keywords: DNA mismatch, DNase I, hydroxyl radical, methidiumpropyl-EDTA-Fe(II)

Identifiers

Local EPrints ID: 56587
URI: https://eprints.soton.ac.uk/id/eprint/56587
ISSN: 1470-8728
PURE UUID: f74b3e1e-ee0e-4275-99e4-d14c6c47c0b0
ORCID for K.R. Fox: ORCID iD orcid.org/0000-0002-2925-7315

Catalogue record

Date deposited: 06 Aug 2008
Last modified: 14 Mar 2019 01:56

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Author: J. Brown
Author: T. Brown
Author: K.R. Fox ORCID iD

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