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#mismatch repair

6 public questions tagged with this topic.

Repair Pathway

MutH introduces a nick in the

MutH is methylation-sensitive endonuclease whose nicking activity is licensed by MutS-MutL complex bound at mismatch. Parental strands carry N6-methyladenine at GATC sites due to Dam methylase, whereas newly synthesized strands remain unmethylated for several minutes post-replication. MutH senses this chemical asymmetry and introduces single-strand break 5' to GATC on unmethylated strand. This incision creates entry point for helicase II UvrD and single-strand exonucleases ExoI, ExoVII, RecJ. Selective cleavage ensures degradation targets error-containing nascent strand while methylated templa

Ref: Berg et al., Biochemistry, 9th ed., Section 28.3: MutH endonuclease hemimethylated incision mechanism; J Biol Chem 2020 strand-specific nicking

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Repair Pathway

Which protein recognizes mismatch in E. coli?

MutS functions as initial mismatch sensor in E. coli mismatch repair pathway. Homodimeric ATPase encircles DNA, conducting diffusive scanning and interrogating duplex for wobble pairing or small insertion-deletion loops that distort helix geometry. Upon encountering mismatch, MutS undergoes ATP-dependent conformational change, kinking DNA about 60 degrees and forming transient sliding clamp. This activated form recruits MutL connector protein, coordinating downstream cleavage and excision steps. Without MutS recognition, proofreading escape errors persist. Homologs MSH2-MSH6 perform analogous

Ref: Lodish et al., Molecular Cell Biology, 9th ed., Chapter 11: MutS mismatch recognition and ATPase; Nature Commun. 2021 MutS clamp loader model

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Repair Pathway

Mismatch repair in E. coli distinguishes strands using

In Escherichia coli, adenine in GATC sequences is methylated by Dam methylase on both strands. Immediately after replication fork passage, parental template remains fully methylated while nascent daughter strand is transiently unmethylated, generating hemimethylated duplexes. MutS scans DNA for mismatches and recruits MutL, which activates latent endonuclease activity of MutH. Recognition of hemimethylated GATC provides temporal strand discrimination signal. MutH selectively nicks unmethylated daughter strand 5' to G, creating entry for UvrD helicase and exonucleases to excise error-containing

Ref: Alberts et al., Molecular Biology of the Cell, 7th ed., Chapter 5: Mismatch repair MutH hemimethylated GATC discrimination; NCBI NBK21279 E. coli MMR mechanism

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Replication Basics

Mismatch repair further improves fidelity after

Despite proofreading, occasional mismatches escape locking before next polymerization step. Post-replicative mismatch repair scans daughter strand discriminated in E. coli by transient hemimethylation of GATC sites remaining unmethylated for minutes. MutS protein detects backbone distortion caused by mismatch, MutL coordinates endonuclease MutH cleavage of unmethylated nascent strand, and helicase plus exonucleases remove error-containing segment. Resynthesis by Pol III restores correct sequence using parental strand template. This pathway improves fidelity additional 100 to 1000-fold and expl

Ref: Alberts et al., Molecular Biology of the Cell, 7th ed., Chapter 5: Methyl-directed mismatch repair after replication

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Restriction Digestion & Mapping

A mismatched site repaired 50% of time will show:

Mismatch repair corrects heteroduplex errors post-replication maintaining genome fidelity. When target site contains mismatch that restores restriction recognition after repair, population outcome depends on repair fidelity and efficiency. Full repair yields uniform cleavable or non-cleavable molecules displaying single digestion pattern. Absence of repair yields opposite parental pattern. When efficiency is about fifty percent, culture contains mixture of repaired and unrepaired molecules in roughly equal proportion. Subsequent restriction digestion then produces composite pattern showing bot

Ref: NCERT Biology Class XII Principles on Klenow fill-in labeling, Lehninger Chapter 9 DNA cloning techniques, and Molecular Cloning by Sambrook Chapter 10 documenting end-labeling of cohesive termini.

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Restriction Digestion & Mapping

In case of 50% efficiency of mismatch repair, what bands are expected after digestion?

When a heteroduplex containing a mismatched restriction site undergoes mismatch repair, outcome depends on repair efficiency and strand bias. With complete repair, all molecules regain the restriction site and show uniform digestion pattern after electrophoresis. With no repair, all retain mismatch and show parental uncut pattern. At fifty percent efficiency, population becomes heterogeneous: half molecules are repaired and cleavable, half remain resistant. Gel then displays both parental and repaired bands simultaneously, reflecting mixed population important for interpreting heteroduplex ana

Ref: NCERT Biology Class XII Principles on Klenow fill-in labeling, Lehninger Chapter 9 DNA cloning techniques, and Molecular Cloning by Sambrook Chapter 10 documenting end-labeling of cohesive termini.

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