The low overall rate of mutation during DNA replication (1 base pair change in
one billion base pairs per replication cycle) does not reflect the true number
of errors that take place during the replication process. The number is kept so
low by a proof-reading system that checks newly synthesized DNA for errors and
corrects them when they are found. Errors in DNA replication can take different
forms, but usually revolve around the addition of a nucleotide with the
incorrect base, meaning the pairing between the parent and daughter strand bases
is not
complementary.
The addition of an
incorrect base can take place by a process called tautomerization. A
tautomer of a base group is a slight rearrangement of its electrons that allows
for different bonding patterns between bases. This can lead to the incorrect
pairing of C with A instead of G, for example.
Figure %: Tautomerization of Cytosine
DNA retains its high level of accuracy is with its proof-reading function.
The 3' to 5' Proof-Reading Exonuclease
The 3' to 5' proof-reading exonuclease works by scanning along directly behind
as the DNA polymerase adds new nucleotides to the growing strand. If the last
nucleotide added is mismatched, then the entire replication holoenzyme backs up,
removes the last incorrect base, and attempts to add the correct base again.
The enzyme is "3' to 5'" because it scans in the opposite direction of DNA
replication, which we learned must always be 5' to 3'. The mechanism of the
proof-reading system offers an explanation as to why DNA replication must occur
in this direction.
Keeping in mind the chemical mechanism we learned for the addition of
nucleotides to the growing DNA strand, imagine what happens when the proof-
reading system removes an incorrectly paired base. The exonuclease removes the
base by cleaving the phosphodiester bond that had just been formed. In 5' to 3'
synthesis, this leaves the 3' -OH still attached to the terminal end of the
growing strand ready to attack another nucleotide.
Figure %: 3' to 5' Exonuclease Action
If synthesis occurred in the opposite direction, the terminal end of a growing
chain would contain a triphosphate group instead of an -OH group. This
triphosphate would become the target of the proof-reading exonuclease and its
removal would halt DNA replication.
Figure %: Incorrect 3' to 5' Synthesis
Types of DNA Damage
After DNA has been completely replicated, the daughter strand is often not a
perfect copy of the parent strand it came from. Mutations during replication
and damage after replication make it necessary for there to be a repair system
to fix any errors in newly synthesized DNA. There are three main sources of
damage to DNA.
Attack by water which can lead to the removal of an amine group from the
base group of a nucleotide or the loss of the entire base group.
Chemical damage that permanently alters the structure of the DNA.
Radiation damage which can lead to nicks in the backbone of DNA or the
formation of thymine dimers, which will be discussed later.
These different sources of damage lead to different categories of DNA damage.
The damage that is caused by water attack can lead to unnatural bases. Chemical
and radiation damage leads to the formation of bulky adducts to, or breaks in,
the growing DNA strand. In the previous section we discussed the 3' to 5'
proof-reading exonuclease that is responsible for fixing mismatches. Because it
is not a perfect system, it can miss mismatched bases. As a result, a third
category of DNA damage is mismatched bases.
