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 language | English (US) |
|---|---|
| Pages (from-to) | 2026-2036 |
| Number of pages | 11 |
| Journal | Nucleic Acids Research |
| Volume | 37 |
| Issue number | 6 |
| DOIs | |
| State | Published - 2009 |
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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 journal › Article
}
TY - JOUR
T1 - A junction branch point adjacent to a DNA backbone nick directs substrate cleavage by Saccharomyces cerevisiae Mus81-Mms4
AU - Ehmsen, Kirk Tevebaugh
AU - Heyer, Wolf Dietrich
PY - 2009
Y1 - 2009
N2 - 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.
AB - 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.
UR - http://www.scopus.com/inward/record.url?scp=64549135730&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=64549135730&partnerID=8YFLogxK
U2 - 10.1093/nar/gkp038
DO - 10.1093/nar/gkp038
M3 - Article
C2 - 19211663
AN - SCOPUS:64549135730
VL - 37
SP - 2026
EP - 2036
JO - Nucleic Acids Research
JF - Nucleic Acids Research
SN - 0305-1048
IS - 6
ER -