Mismatch Repair

Core Concept

Mismatch repair (MMR) is a post-replicative system that corrects base–base mismatches and small insertion/deletion loops that escape the polymerase's proofreading exonuclease. By specifically excising and resynthesizing a stretch of the newly synthesized strand, MMR lowers the overall replication error rate by a further ~100–1000-fold, to roughly one error per 10⁹–10¹⁰ base pairs. Loss of MMR causes a "mutator" phenotype and is the molecular basis of hereditary nonpolyposis colorectal cancer (Lynch syndrome).

Key Components

• MutS homologs (MSH2–MSH6 or MSH2–MSH3) — recognize mismatches and insertion/deletion loops.
• MutL homologs (MLH1–PMS2) — couple mismatch recognition to downstream excision.
• Exonuclease 1 (EXO1) — removes the mismatched strand segment.
• Proliferating cell nuclear antigen (PCNA) — directs the machinery to the correct strand and coordinates resynthesis.
• Replication protein A (RPA), DNA polymerase δ, and DNA ligase — complete gap filling and sealing.

Mechanism

MMR exploits the fact that, immediately after replication, the newly synthesized strand is distinguishable from the template. In bacteria this is done by transient hemimethylation (the new strand is unmethylated); in eukaryotes the strand-discrimination signal is linked to the replication machinery — nicks and PCNA mark the nascent strand. The MutS complex binds the mismatch, recruits MutL, and together they direct EXO1 to nick and degrade the error-containing strand past the mismatch; DNA polymerase δ then resynthesizes the excised region and ligase seals it.

How It Works

1. A mismatch (e.g., G paired with T) or a small loop persists after replication because proofreading missed it.
2. The MSH2–MSH6 heterodimer recognizes and binds the distortion.
3. MLH1–PMS2 is recruited, forming a complex that scans for the strand-discrimination signal (the nick/PCNA on the nascent strand).
4. EXO1 is directed to the nascent strand and digests it from the nick through and beyond the mismatch.
5. RPA stabilizes the resulting single-stranded gap; DNA polymerase δ fills it using the intact template.
6. DNA ligase seals the final nick, restoring a correct, fully paired duplex.

Energy and Directionality

Excision and resynthesis are ATP- and dNTP-consuming. MutS and MutL are ATPases whose nucleotide binding/hydrolysis drives conformational changes and the long-range communication between mismatch and strand signal. Directionality is provided by the strand-discrimination signal — the mismatch is removed only from the newly synthesized strand, so the parental (correct) sequence is always preserved. The resynthesis step is powered by dNTP hydrolysis, as in replication.

Experimental Evidence

• Bacterial genetics — mutator strains (mutS, mutL) accumulate point mutations at ~100–1000× the normal rate, proving the genes correct replication errors.
• In vitro reconstitution — purified MutS, MutL, EXO1, PCNA, pol δ, and ligase carry out nick-directed mismatch excision, defining the eukaryotic pathway.
• Microsatellite instability (MSI) — MMR-defective cells show length instability of short repeats, a diagnostic hallmark used in cancer screening.
• Lynch syndrome linkage — germline mutations in MSH2, MLH1, MSH6, or PMS2 cause hereditary colorectal and other cancers, directly linking MMR to tumor suppression.

Technique

Detection of MMR status uses microsatellite instability testing (PCR of repeat loci) and immunohistochemistry for MLH1/MSH2/MSH6/PMS2 on tumors. Functional assays include in vitro mismatch repair of heteroduplex DNA and mutation-accumulation (mutator) assays. Genome-wide mutation spectra are read by whole-genome sequencing.

How it works

1. A mismatch (e.g., G paired with T) or a small loop persists after replication because proofreading missed it.
2. The MSH2–MSH6 heterodimer recognizes and binds the distortion.
3. MLH1–PMS2 is recruited, forming a complex that scans for the strand-discrimination signal (the nick/PCNA on the nascent strand).
4. EXO1 is directed to the nascent strand and digests it from the nick through and beyond the mismatch.
5. RPA stabilizes the resulting single-stranded gap; DNA polymerase δ fills it using the intact template.
6. DNA ligase seals the final nick, restoring a correct, fully paired duplex.

Common confusions

• "MMR and proofreading are the same" — proofreading (3′→5′ exonuclease of the polymerase) acts during synthesis; MMR acts after synthesis to catch what proofreading missed.
• "MMR repairs any DNA damage" — it targets replication misincorporation errors; chemically damaged bases are handled by BER/NER.
• "Both strands are at risk of excision" — MMR specifically removes the new strand so the template is preserved.
• "MMR defects only cause colon cancer" — Lynch syndrome also raises risk of endometrial, ovarian, gastric, and other cancers.
• "Microsatellite instability is benign" — it is a diagnostic marker of defective MMR and a cancer driver.

Quick review

• MMR fixes replication mismatches/small loops after synthesis.
• MSH2–MSH6 recognizes; MLH1–PMS2 couples to excision; EXO1 digests the new strand.
• Strand discrimination by nicks/PCNA (eukaryotes) or hemimethylation (prokaryotes).
• Pol δ resynthesizes; ligase seals; ATP and dNTPs power the reaction.
• Defects → MSI and Lynch syndrome.
• Tested by MSI PCR and MLH1/MSH2 immunohistochemistry.