The vsr gene product of E. coli K-12 is a strand- and sequence-specific DNA mismatch endonuclease

Hennecke, Frank; Kolmar, Harald; Bründl, Kerstin; Fritz, Hans-Joachim

doi:10.1038/353776a0

Cited by 133 publications

(91 citation statements)

References 25 publications

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“…: 313-577-2547; Fax: 313-577-8822; E-mail: axb@chem.wayne.edu. and the cell pellets were resuspended in 1 ml of a buffer containing 5 mM CaCl 2 and 10 mM MgSO 4 . One hundred l of the suspensions were infected with 10, 50, or 100 l of the P1 phage, and the cultures were incubated at 37°C for 30 min without shaking.…”

Section: Methodsmentioning

confidence: 99%

“…In E. coli, a specialized mismatch correction process called very short patch repair corrects these mispairs to C⅐G (3). The key enzyme in this repair pathway is a sequence-specific, mismatch-specific endonuclease, Vsr, which hydrolyzes the phosphodiester linkage preceding the mismatched T (4). No eukaryotic sequence homologs of this enzyme have been reported; instead a DNA glycosylase is thought to serve the same function (5).…”

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confidence: 99%

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The Role of the Escherichia coli Mug Protein in the Removal of Uracil and 3,N 4-Ethenocytosine from DNA

Lutsenko

Bhagwat

1999

Journal of Biological Chemistry

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The human thymine-DNA glycosylase has a sequence homolog in Escherichia coli that is described to excise uracils from U⅐G mismatches (Gallinari, P., and Jiricny, J. (1996) Nature 383, 735-738) and is named mismatched uracil glycosylase (Mug). It has also been described to remove 3,N 4 -ethenocytosine (⑀C) from ⑀C⅐G mismatches (Saparbaev, M., and Laval, J. (1998) Proc. Natl. Acad. Sci. U. S. A. 95, 8508 -8513). We used a mug mutant to clarify the role of this protein in DNA repair and mutation avoidance. We find that inactivation of mug has no effect on C to T or 5-methylcytosine to T mutations in E. coli and that this contrasts with the effect of ung defect on C to T mutations and of vsr defect on 5-methylcytosine to T mutations. Even under conditions where it is overproduced in cells, Mug has little effect on the frequency of C to T mutations. Because uracil-DNA glycosylase (Ung) and Vsr are known to repair U⅐G and T⅐G mismatches, respectively, we conclude that Mug does not repair U⅐G or T⅐G mismatches in vivo. A defect in mug also has little effect on forward mutations, suggesting that Mug does not play a role in avoiding mutations due to endogenous damage to DNA in growing E. coli. Cell-free extracts from mug ؉ ung cells show very little ability to remove uracil from DNA, but can excise ⑀C. The latter activity is missing in extracts from mug cells, suggesting that Mug may be the only enzyme in E. coli that can remove this mutagenic adduct. Thus, the principal role of Mug in E. coli may be to help repair damage to DNA caused by exogenous chemical agents such as chloroacetaldehyde.Cytosine is the most unstable of the four bases in DNA and deaminates hydrolytically to create U⅐G mismatches. If unrepaired, uracil can pair with an adenine during replication causing a C to T mutation. For this reason, cells contain uracil-DNA glycosylase (Ung), an enzyme that removes the uracil and initiates its replacement with cytosine. The importance of Ung in mutation avoidance is evidenced by the observation that ung strains of Escherichia coli (1) and yeast (2) accumulate C to T mutations.Cytosines methylated at position 5 similarly deaminate to create T⅐G mismatches, which are not subject to repair by Ung. In E. coli, a specialized mismatch correction process called very short patch repair corrects these mispairs to C⅐G (3). The key enzyme in this repair pathway is a sequence-specific, mismatch-specific endonuclease, Vsr, which hydrolyzes the phosphodiester linkage preceding the mismatched T (4). No eukaryotic sequence homologs of this enzyme have been reported; instead a DNA glycosylase is thought to serve the same function (5). This enzyme excises thymines from T⅐G mismatches (6) and prefers mismatches that are followed by a G⅐C pairs (7,8). This enzyme, thymine-DNA glycosylase (TDG), 1 could prevent mutations when 5-methylcytosines within CG dinucleotides deaminate to thymine.The cDNA for TDG was cloned and its sequence was determined (9). Remarkably, a sequence homolog of this protein was found in E. coli and Serratia ma...

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Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

The Role of the Escherichia coli Mug Protein in the Removal of Uracil and 3,N 4-Ethenocytosine from DNA

Lutsenko

Bhagwat

1999

Journal of Biological Chemistry

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“…In the case of VSPR, the MutS and MutL proteins recruit the Vsr endonuclease, which creates a single-stranded nick adjacent to the mismatched thymine on its 5′ side [23]. Pol I then removes and replaces a small number of bases (<10) 3′ to the nick utilizing its 5′ → 3′ exonuclease and polymerase activities [24].…”

Section: Introductionmentioning

confidence: 99%

Mismatch repair proteins collaborate with methyltransferases in the repair of O6-methylguanine

et al. 2008

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DNA repair is essential for combatting the adverse effects of damage to the genome. One example of base damage is O 6 -methylguanine (O 6 mG), which stably pairs with thymine during replication and thereby creates a promutagenic O 6 mG:T mismatch. This mismatch has also been linked with cellular toxicity. Therefore, in the absence of repair, O 6 mG:T mismatches can lead to cell death or result in G:C→A:T transition mutations upon the next round of replication. Cysteine thiolate residues on the Ada and Ogt methyltransferase (MTase) proteins directly reverse the O 6 mG base damage to yield guanine. When a cytosine is opposite the lesion, MTase repair restores a normal G:C pairing. However, if replication past the lesion has produced an O 6 mG:T mismatch, MTase conversion to a G:T mispair must still undergo correction to avoid mutation. Two mismatch repair pathways in E. coli that convert G:T mispairs to native G:C pairings are methyl-directed mismatch repair (MMR) and very short patch repair (VSPR). This work examined the possible roles that proteins in these pathways play in coordination with the canonical MTase repair of O 6 mG:T mismatches. The possibility of this repair network was analyzed by probing the efficiency of MTase repair of a single O 6 mG residue in cells deficient in individual mismatch repair proteins (Dam, MutH, MutS, MutL, or Vsr). We found that MTase repair in cells deficient in Dam or MutH showed wild-type levels of MTase repair. In contrast, cells lacking any of the VSPR proteins MutS, MutL, or Vsr showed a decrease in repair of O 6 mG by the Ada and Ogt MTases. Evidence is presented that the VSPR pathway positively influences MTase repair of O 6 mG:T mismatches, and assists the efficiency of restoring these mismatches to native G:C base pairs.

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“…The T:G mismatch generated by spontaneous deamination of the 5-mC is repaired in E. coli by a specific pathway that is able to discriminate between the erroneous thymidine and the correct guanine and selectively remove the incorrect thymidine (22). This repair mechanism is known as very short patch (VSP) repair reflecting the short (often less than 10 bases) patch used to restore the correct DNA sequence (23).…”

mentioning

confidence: 99%

“…The gene product of vsr, the Vsr endonuclease, catalyzes the initiating step in VSP repair. The Vsr endonuclease recognizes the T:G mismatch in the appropriate sequence context and cleaves 5Ј to the mismatched T, leaving a nick with a 3Ј-OH and a 5Ј-PO 4 (22,(31)(32)(33)(34). DNA polymerase I is thought to bind at the nick produced by Vsr cleavage and undergo nick translates, using its 5Ј to 3Ј exonuclease activity and its 5Ј to 3Ј polymerase activity, restoring the correct C:G base pair while adding a minimal number of additional nucleotides (27).…”

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confidence: 99%

Reconstitution of the Very Short Patch Repair Pathway from Escherichia coli

Robertson

Matson

2012

Journal of Biological Chemistry

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Background: Very short patch repair rectifies thymidine-guanine mismatches arising from spontaneous deamination of 5-methyl cytosine. Results: The very short patch repair pathway was reconstituted using purified enzymes. Conclusion: Patch length depends on the concentration of DNA ligase and may be regulated by the addition of MutS and MutL. Significance: The results suggest roles for MutL and MutS in very short patch repair.

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The vsr gene product of E. coli K-12 is a strand- and sequence-specific DNA mismatch endonuclease

Cited by 133 publications

References 25 publications

The Role of the Escherichia coli Mug Protein in the Removal of Uracil and 3,N 4-Ethenocytosine from DNA

The Role of the Escherichia coli Mug Protein in the Removal of Uracil and 3,N 4-Ethenocytosine from DNA

Mismatch repair proteins collaborate with methyltransferases in the repair of O6-methylguanine

Reconstitution of the Very Short Patch Repair Pathway from Escherichia coli

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