Abstract
We have developed a purified system that supports mismatch-dependent 5'-->3' excision. In the presence of RPA, ATP, and a mismatch, MutSalpha activates 5'-->3' excision by EXOI, and excision terminates after removal of the mispair. MutSalpha confers high processivity on EXOI, and termination is due to RPA-dependent displacement of this processive complex from the helix and a weak ability of EXOI to reload at the RPA-bound gap in the product, as well as MutSalpha- and MutLalpha-dependent suppression of EXOI activity in the absence of a mismatch cofactor. As observed in the purified system, excision directed by a 5' strand break in HeLa nuclear extract can proceed in the absence of MutLalpha or PCNA, although 3' excision in the extract system requires both proteins.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Adenosine Triphosphatases / metabolism
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Adenosine Triphosphate / metabolism
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Bacterial Proteins*
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Base Pair Mismatch*
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Cell Nucleus / chemistry
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DNA / genetics
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DNA / metabolism*
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DNA Repair Enzymes
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DNA Repair*
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DNA-Binding Proteins / metabolism
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Enzyme Activation
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Escherichia coli Proteins / metabolism
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Exodeoxyribonucleases / metabolism
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HeLa Cells
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Humans
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MutL Proteins
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MutS DNA Mismatch-Binding Protein
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Replication Protein A
Substances
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Bacterial Proteins
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DNA-Binding Proteins
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Escherichia coli Proteins
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MutL protein, E coli
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RPA1 protein, human
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Replication Protein A
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Adenosine Triphosphate
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DNA
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EXO1 protein, human
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Exodeoxyribonucleases
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exodeoxyribonuclease I
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Adenosine Triphosphatases
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MutL Proteins
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MutS DNA Mismatch-Binding Protein
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MutS protein, E coli
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DNA Repair Enzymes