Molecular Biology

Molecular Genetics

mismatch repair

Most mismatches are caused by replication errors. However, mismatches can also be produced by other mechanisms, such as deamination of 5-methyl cytosine to generate T improperly paired to G. Where the appropriate DNA-N-glycosylase is available, mismatches can also be repaired by base excision repair.

Mismatch repair has been studied most extensively in E. coli, where the proteins MutS, MutL, and MutH initiate the repair process. Newly synthesized strands are not immediately methylated in E.coli. First, MutS recognizes and binds to true mismatches and insertions/deletions of up to 4 nucleotides. Next, MutL binds to and stabilizes this complex of MutS/mismatched strand. The MutS-MutL complex then activates MutH, which locates a nearby methyl group and creates a nick in the newly synthesized strand opposite the methyl group. Excision is accomplished in E.coli by cooperation between the UvrD (Helicase II) protein, which unwinds from the nick toward the mismatch, and a single-strand specific exonuclease of appropriate polarity. Finally, resynthesis by Polymerase III and ligation by a DNA ligase repair the sequence and re-ligate the strands.

Unlike the un-methylated new strands in E.coli, strand-specificity in eukaryotes may be signalled by single-strand nicks. In nascent eukaryotic DNA strands, single-strand breaks occur between Okazaki fragments in the lagging strand and at the 3' end of the leading strand.

Eukaryotes lack homologues of MutH and uvrD, but do possess numerous homologues of MutS and MutL (MSHs 1-6, MLHs 1-3 and PMS 1 or 2). In E.coli, MutS and MutL function as monomers. The homologous eukaryotic proteins function as heterodimers. Human cells also possess two heterodimers of MutS homologues – MutSalpha (MSH2/MSH6), which recognizes single base mismatches and small loops, and MutSbeta (MSH2/MSH3), which recognizes small loops.

In eukaryotic cells, several standard replication proteins are needed for mismatch repair. The "clamp" protein, PCNA is a cofactor for most DNA polymerases and stabilizes the MutS and MutL heterodimers at mismatch sites on DNA. Three MutL homolog dimers are also known – in humans, MLH1/PMS2, MLH1/PMS1, and MLH1/MLH3. Further, just as exonucleases are thought to be important for mismatch repair in prokaryotes, at least two nucleases appear to contribute to mismatch repair in eukaryotic cells – exonuclease 1 and Flap Endonuclease (FEN-1 or DNase IV; Rad27 in S. cerevisiae). PCNA is also required during the later DNA synthesis step of mismatch repair. The DNA synthesis step also requires RPA (the eukaryotic single-stranded DNA-binding protein), Replication factor C (which loads PCNA onto DNA molecules at primer termini) and DNA polymerase delta.

Hereditary non-polyposis colon cancer (HNPCC) is a form of colon cancer frequently associated with defects in the genes encoding MSH2 (about 35% of identified gene-defect cases) and MLH1 (about 60% of identified gene-defect cases). HNPCC is characterized by early age of onset and autosomal dominant inheritance with high penetrance.

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