A junction branch point adjacent to a DNA backbone nick directs substrate cleavage by Saccharomyces cerevisiae Mus81-Mms4

Kirk Tevebaugh Ehmsen, Wolf Dietrich Heyer

Research output: Contribution to journalArticle

21 Citations (Scopus)

Abstract

The DNA structure-selective endonuclease Mus81-Mms4/Eme1 incises a number of nicked joint molecule substrates in vitro. 3′-flaps are an excellent in vitro substrate for Mus81-Mms4/Eme1. Mutants in MUS81 are synthetically lethal with mutations in the 5′-flap endonuclease FEN1/Rad27 in Saccharomyces cerevisiae and Schizosaccharomyces pombe. Considering the possibility for isoenergetic interconversion between 3′- and 5′- flaps, these data are consistent with the hypothesis that Mus81-Mms4/Eme1 acts on 3′-flaps in vivo. FEN1/Rad27 prefers dually flapped substrates and cleaves in a way that allows direct ligation of the resulting nick in the product duplex. Here we test the activity of Mus81-Mms4 on dually flapped substrates and find that in contrast to FEN1/Rad27, Mus81-Mms4 activity is impaired on such substrates, resulting in cleavage products that do not allow direct religation. We conclude that Mus81-Mms4, unlike FEN1/Rad27, does not prefer dually flapped substrates and is unlikely to function as a 3′-flapase counterpart to the 5′-flapase activity of FEN1/Rad27. We further find that joint molecule incision by Mus81-Mms4 occurs in a fashion determined by the branch point, regardless of the position of an upstream duplex end. These findings underscore the significance of a nick adjacent to a branch point for Mus81-Mms4 incision.

Original languageEnglish (US)
Pages (from-to)2026-2036
Number of pages11
JournalNucleic Acids Research
Volume37
Issue number6
DOIs
StatePublished - 2009

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Single-Stranded DNA Breaks
Saccharomyces cerevisiae
Flap Endonucleases
Joints
Schizosaccharomyces
Endonucleases
Ligation
Mutation
DNA
In Vitro Techniques

ASJC Scopus subject areas

  • Genetics

Cite this

A junction branch point adjacent to a DNA backbone nick directs substrate cleavage by Saccharomyces cerevisiae Mus81-Mms4. / Ehmsen, Kirk Tevebaugh; Heyer, Wolf Dietrich.

In: Nucleic Acids Research, Vol. 37, No. 6, 2009, p. 2026-2036.

Research output: Contribution to journalArticle

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