Chapter 16 - Repair Systems Flashcards
Direct Repair
LEAST COMMON. Reversal/Removal of the damage.
Excision Repair
One strand of DNA is directly excised and then replaced by resynthesis using the complementary strand as template.
Mismatch Repair
MOST COMMON. Corrects recently inserted bases that do not pair properly.
Recombination Repair
Filling a gap in one strand of duplex DNA by retrieving a homologous single strand from another duplex.
Nonhomologous end joining
Rejoins broken DSB.
Deamination of cytosine
- Creates a U-G base pair.
- Uracil is preferentially removed from the mismatched pair.
- Minor structural distortion with pyrimidine-purine pair (U-G).
Replication Error
- Causes a mismatched pair.
- Corrected by replacing one base.
- Greater structural distortion with purine-purine pair (A-G).
- If uncorrected, a mutation is fixed in one daughter duplex.
Ultraviolet irradiation
- Thymine dimer formation.
- Corrected by excision.
- The dimer blocks replication and transcription.
Methylation of the Base
- Causes mispairing at replication.
* Corrected by dealkylation.
Depurination
- Removes a base from DNA.
- Corrected by insertion
- Blocking replication and transcription.
Nucleotide excision repair removes
a stretch of DNA that includes the damaged base.
Incision
Requires the activity of an endonuclease to recognize and cut on both sides of the damaged DNA.
Distinguishes nucleotide from Base Excision Repair.
Excision & Synthesis
Removal and replacement often occur concurrently and
require an exonuclease/helicase (excision) and DNA polymerase (synthesis).
Ligation of Nicks
Requires DNA ligase to covalently link the 3’-ends of the new DNA strand with the original DNA.
Uvr repair system
In E. coli, the Uvr repair system accounts for almost all of the excision repair events. Uvr complex can be directed to sites of damage by other proteins. 1. UvrAB dimer recognizes and binds damaged DNA. 2. UvrA released (ATP-dependent) and UvrC binds. 3. UvrC nicks 10-11 bases around the damage site; ATP-dependent (Incision). 4. UvrD is the helicase that unwinds the DNA to release ssDNA strand.
UvrAB
Dimer recognizes and binds damaged DNA.
UvrA
UvrA released (ATP-dependent)
UvrC
Nicks 10-11 bases around the damage site; ATP-dependent
Incision
UvrD
Helicase that unwinds the DNA to release ssDNA strand.
Mfd
Recognizes a stalled RNA polymerase (transcription) and
directs the repair complexes to the damaged template strand.
Displaces the ternary RNA polymerase from the DNA.
Promotes UvrA/B recruitment.
Human excision repair mechanism
In eukaryotes, a complex of proteins including XP products and the transcription factor TFIIH.
Nucleotide excision repair occurs via 2 major pathways:
- Global Genome Repair
2. Transcription-coupled Repair
Global Genome Repair
Recognizes damage anywhere in the genome.
Transcription-coupled Repair
Preferentially repairs transcriptionally active genes.
Shared components of the 2 pathways (Global Genome Repair, Transcription-coupled Repair) include:
- Transcription factor TFIIH
- XPB and XPD
- XPF and XPG
- DNA synthesis
- Nicks are ligated via DNA ligase II and XRCC1.
XPB and XPD
Helicase components; exhibit activity to unwind around
the damaged site in Global Genome Repair, Transcription-coupled Repair.
XPF
5’ exonuclease for incision around the damaged base in Global Genome Repair, Transcription-coupled Repair.
XPG
3’ exonuclease for incision around the damaged base in Global Genome Repair, Transcription-coupled Repair.
DNA ligase II and XRCC1
Ligate nicks in Global Genome Repair, Transcription-coupled Repair.
Base excision repair
Removes the individual damaged base from DNA by excision and then re-synthesis (not “flipping”).
Base removal triggers the removal and replacement of a stretch of polynucleotides.
The nature of the base removal reaction determines which of two pathways for excision repair is activated.
Glycosylase
Hydrolyzes the bond between base and deoxyribose (using H2O).
Lyase
Take the reaction further by opening the sugar ring (using NH2).
Attack of the deoxyribose ring recruits the polβ pathway to replace a short stretch.
Trigger for mammalian excision-repair pathways
Single base removal by glycosylase or lyase action.
Examples of enzymes that remove/modify an individual base from DNA by “flipping” the base directly out of the
DNA duplex.
Alkyladenine DNA glycosylase for alkylated bases (methylated).
Uracil-DNA glycosylase.
Photolyase for pyrimidine dimers.
Yeast Rad4 - flips 2-A bases complementary to T-dimers.
Damaged DNA that has not been repaired
Causes DNA polymerase III to stall during replication.
Prokaryotes DNA polymerase V and
IV or eukaryotic DNA polymerases Eta
and Zeta
Can synthesize a complement to a damaged strand in order to bypass a lesion.
Replication polymerase
Stalls at the site of damage; is displaced by a trans-lesion polymerase; commences DNA synthesis.
MutS
Recognizes a mismatch and translocates to a GATC site.
Bound to both the mismatch site and to DNA as it
translocates, and as a result it creates a loop in the DNA.
Endonucleases degrade the strand from the GATC to the mismatch site.
MutH
Cleaves the un-methylated strand at the GATC.
MutS/L systems
Initiates repair of mismatches in eukaryotic systems produced by replication slippage. Do not use DNA methylation to select the daughter strand for repair. Not fully understood how eukaryotes recognize the daughter
strand during mismatch repair.
Replication fork stalls when it encounters a damaged site/nick in DNA will:
- May reverse by pairing between the two newly synthesized strands.
- May restart after repairing the damage and use a helicase to move the fork forward.
- May initiate translesion/excision repair or recombination repair to correct DNA damage.
Recombination repair
Preferred mechanism for repairing DSB.
Works analogous to Homologous Recombination in Meiotic
cells. Homologous recombination ensures no genetic
information is lost from a broken DNA end.
Double-strand breaks
One of the most severe types of DNA damage that can occur, particularly in eukaryotes.
Can be caused by ionizing radiation, oxygen radicals generated by cellular metabolism, or action of endonuclease.
Ku70 and Ku80
Recognize broken ends by Ku70/Ku80 heterodimer and formation of a scaffold.
An unknown DNA polymerase fills in any remaining ssDNA protrusions.
DNA-dependent protein kinase (DNA-PK)
Activated by DNA to phosphorylate protein targets like
Artemis, which exhibits nuclease activity to trim overhanging ends.
What different mechanisms of recognition do global genome repair and transcription-coupled repair use? (XPC vs. RNA polymerase II).
Global genome repair - XPC
Transcription coupled repair - RNAP II
Glycosylase-mediated APE1 recruitment initiates
The polδ/ε pathway to replace a long polynucleotide stretch.
