Question

During mismatch repair, how does the cell “know” which strand to degrade and repair? Why will it lose this ability after a certain amount of time? Provide a reason that might explain why it is safer to degrade the new strand of DNA rather than the parental strand.

Answer 1

Associated with the replication origin (OriC) of E. coli, there are many copies of 5' GATC 3'; a short palindrome which is the target site for Dam methylase of E.coli. The A residues on both the strands in the Dam target can be methylated and the bacteria always keeps its Dam target sites fully methylated. Replication of the fully methylated DNA forms hemimethylted DNA in which only the A residues in the parental strand have methyl groups and the A residues in the Dam target sites in the newly synthesized strand do not have methyl groups. By identifying the methylated strand bacteria can distinguish the parental strand from the newly synthesized strand. After replication the hemimethylated DNA is converted into the fully methylated condition by completion of methylation of Dam target sites in the newly synthesized strand. The completion of methylation of the Dam target sites associated with the OriC occur after some 12-13 minutes, whereas the Dam target sites elsewhere in the genome is fully methylated immediately after replication. Mismatch correction is made within this 12 to 13 minutes. During this period the bacteria can distinguish the parental strand from the newly synthesized strand. Mismatch correction is made by excising a short stretch of DNA containing the mismatch from the newly synthesized strand and a replacement is made by using the original parental strand as the template. If the parental strand is excised for mismatch correction, then the correction is made using the newly synthesized strand as the template. This results in the occurrence of a point mutathion. The following is an example

Original DNA sequence -   ATAGC

                                           TATCG

Replication error during leading to a mismatch

                                           ATAGC

                                           TATCG   Normal daughter

                                           ATAGC

                                           TATTG   Daughter with a mismatch

Mismatch correction by excising the newly synthesized strand

                                           ATAGC

                                           TATCG

Mismatch correction by excising the parental strand

                                          ATAAC

                                           TATTG

The original G-C pair is convered into an A-T pair.

Correction should be made within 12-13 minutes, when the Dam target sites associated with the OriC is awaiting methylation. When this methylation is completed it is not possible to distinguish the parental strand from the newlly made strand.