DNA Errors and Repair
Replication Fidelity
With all of the enzymes of replication flying around it seems like our newly synthesized strand would be filled with errors. This couldn’t be farther from the truth though. DNA polymerase only makes 1 error per 100,000 bases. Even still our genomes are so large, around 6 billion base pairs, that this low error rate adds up to 120,000 errors per division. However, the final replicated strand ends up containing far fewer errors. What accounts for the difference between the actual error rate and the DNA polymerase error rate then?
Proofreading
DNA polymerase is extremely good at checking its work in a process called proofreading. Even if it places down the wrong base pair the enzyme will later double-check its work and detect mismatched base pairs. It then excises them and replaces them with the correct base pair and checks its work again just to be sure.
How does DNA polymerase know which strand to trust though? Couldn’t it start changing the parent strand if it accidentally thought the newly synthesized daughter strand was the correct template? It could if the two strands were identical but they aren’t. While we hope the base pairs in DNA are an exact copy of the parent strand, DNA also accumulates modifications throughout its lifespan. One of the most common is methylation. So the older the strand the more methylation it has. DNA polymerase recognizes this and uses the older more methylated strand as the “correct” strand since it represents the parent strand.
Mismatch Repair
Even still proofreading isn’t perfect and leaves errors in place. Thankfully cells have another mechanism to make sure these errors don’t end up in the final DNA strand, mismatch repair. Mismatch repair (MMR) is exactly what it sound like. It fixes mismatched base pairs by detecting base pairing errors, removing the incorrect base, and replacing it with the correct one. By this point our replicated DNA will only have an error every 10,000,000,000 base pairs which is larger than our genome.
DNA Damage and Repair
Now our newly synthesized DNA strand is a perfect copy of the original parent strand however as a cell goes about its daily life it can acquire new DNA errors. These errors occur as the DNA is damaged by normal metabolic processes, sunlight, or physical damage. If left unfixed the DNA of our parent strands would be riddled with errors. In order to deal with DNA damage, cells employ two major repair mechanisms that fix two of the most common types of DNA damage.
Nucleotide Excision Repair
The first is the nucleotide excision repair (NER) mechanism that fixes thymine dimers. Thymine dimers form when the UV component of sunlight hits our DNA strand. The sunlight catalyzes a photochemical reaction that results in the bonding of adjacent thymine molecules. When this occurs the DNA backbone bulges outwards causing a disruption in the normal smooth helical structure of DNA.
NER recognizes this bulge and cuts out the distorted base pairs plus some by cutting through the phosphodiester backbone. Since the DNA backbone was formed by the generation of new P-O bonds, the NER will nick open these same P-O bonds. Then DNA polymerase can come along and replace the gap with new thymine residues that are bonded to their base-pairing adenines rather than one another.
The enzyme that facilitates this process is known as an endonuclease. The endo refers to inside and the nuclease refers to the fact that these are hydrolytic enzymes that cut apart nucleotides. Therefore an endonuclease cleaves the nucleotides found in the middle of the DNA strand.
This is in contrast to exonucleases which remove nucleotides from the end of a DNA strand. Since exo means outside we can remember that an exonuclease will cut the outside of the DNA strand i.e. the ends.
Base Excision Repair
Lastly, other types of DNA damage can result in the switching of one nitrogenous base to another. For example one of the cytosine’s amino groups can be removed in a process called deamination. When this occurs cytosine is converted into uracil and no longer base pairs with guanine. Instead, the uracil will now pair with adenine so if this DNA strand underwent replication prior to repair the new strand would have a mutated base pair.
To fix this our cells use base excision repair (BER). BER works in an identical fashion to the NER except instead of cutting out a chunk of DNA the endonucleases involved here only cut the single incorrect base pair out. Again DNA polymerase then fills the gap and DNA ligase joins the backbone together again.