Chapter 25.2: DNA Repair

• DNA molecules are irreplaceable.
• Damaged proteins and RNA molecules can be replaced using information encoded in the DNA.
• Mutations are linked to cancer.
• A mutation is a permanent change in the nucleotide sequence.
• A substitution mutation is the replacement of one base pair with another.
• An insertion mutation is the addition of 1 or more base pairs.
• A deletion mutation is the deletion of one or more base pairs.
• A silent mutation is a mutation that affects nonessential DNA or has a negligible effect on gene function.
• Mutations rarely confer some biological advantage.
• Most nonsilent mutations are neutral or deleterious.
• The Ames test measures the potential of a given chemical compound to promote certain easily detected mutations in a strain of bacteria that cannot synthesize histidine. Strains that grow contain mutations that restore the ability to synthesize histidine.
• Nearly 200 genes in the human genome encode proteins dedicated to DNA repair.
• Many DNA repair processes seem to be extraordinarily inefficient energetically. This is irrelevant because the integrity of the genetic information is more important.
• Mismatches are corrected to reflect the template strand information.
• The template strand is distinguished from the newly synthesized strand by the presence of methyl group tags on the template DNA.
• In E. coli, Dam methylase methylates DNA at the N6 position of adenines within the 5'-GATC sequences.
• In methyl-directed mismatch repair, MutS recognices the mismatch. MutH then recognizes the 5'-GATC sequence and has site specific endonuclease activity. MutL binds both MutS and MutH. MutSL complex forms around the DNA at the mismatch and moves along the DNA to MutH bound at the hemimethylated 5'-GATC. MutH cleaves the unmethylated strand on the 5' side of G in the GATC sequence.
• In methyl directed repair, the unmethylated strand is unwound and degraded in the 3' to 5' direction from cleavage site through the mismatch.
• In methyl directed repair, the cleaved segment is replaced with new DNA by SSB, exonuclease I, X, and VII, DNA polymerase III, and DNA ligase.
• Mismatch repair on the 3' side of the cleavage site in methyl directed repair is similar to mismatches on the 5' side. The exonuclease is either exonuclease VII or RecJ nuclease.
• Mismatch repair is particularly costly for E. coli.
• Mismatches may occur over 1000 base pairs from the GATC sequence. Repair requires degradation and replacement of a strand segment of this entire length.
• Eukaryotic mismatch repair systems have several proteins that are structurally and functionally analogous to bacterial MutS and MutL proteins.
• Many details of eukaryotic mismatch repair are unknown.
• In eukaryotic mismatch repair systems, identification of newly synthesized DNA strands does not involve GATC sequences.
• DNA glycosylases recognize common DNA lesions and remove the affected base by cleaving the N-glycosyl bond in the process of base excision repair. It is generally specific for one lesion type.
• The AP or abasic site is an apurinic or apyrimidinic site in the DNA resulting from the removal of a base by DNA glycosylase.
• Uracil DNA glycosylases specifically remove from DNA the uracil that results from spontaneous deamination of cytosine.
• Uracil DNA glycosylases do not remove uracil from RNA or thymine residues from DNA.
• Deamination is 100 fold faster in ssDNA.
• Other DNA glycosylases recognize formamidopyrimidine (from purine oxidation), 8-hydroxyguanine (from purine oxidation), hypoxanthine (from adenine deamination), alkylated bases, and pyrimidine dimers.
• In bacteria in base excision repair, the deoxyribose 5'-phosophate left behind is removed and replaced with a new nucleotide.
• In bacteria undergoing base excision repair, AP exonucleases cut the DNA strand containing the AP site, DNA polymerase I replaces DNA, and DNA ligase seals the remaining nick.
• The nucleotide excision system repairs DNA lesions that cause large distortions in the DNA helical structure.
• Exinuclease is a multi subunit enzyme that hydrolyzes two phosphodiester bonds, one on either side of the distorsion.
• Nucleotide excision repair involves excinuclease, DNA polymerase I or DNA polymerase epsilon fills in the gap, and DNA ligase seals the nick.
• The ABC excinuclease has three components, UvrA, UvrB, and UvrC.
• UvrA is a part of the ABC excinuclease that is a dimeric ATPase that scans DNA and binds to the lesion site.
• UvrB of the ABC excinuclease binds to UvrA and makes an incision at the fiftth phosphodiester bond on the 3' side of the lesion after UvrA dissociates from the lesion.
• UvrC of the ABC excinuclease binds UvrB and makes an incision at the eighth phosphodiester bond on the 5' side of the lesion after UvrB does.