Spontaneous alterations likely to require DNA repair

The sites on each nucleotide that are known to be modified by spontaneous oxidative damage (red
arrows) hydrolytic attack (blue arrows) and uncontrolled methylation by the methyl group donor Sadenosylmethoinine (green arrows) are shown with the width of each arrow indicating the relative
frequency of each event. ( After T. Lindahl Nature 362:709-715 1993 With permission from
Macmillan Publisher Ltd.)

Depurination and Deamination

Depurination

Deamination

These two reactions are the most frequent spontaneous chemical reactions known to
create serious DNA damage in cells. Depurination can release guanine (shown here) as
well as adenine from DNA. The major type of deamination reaction converts cytosine to
an altered DNA base uracil (shown here) but deamination occurs on other bases as well.
These reactions normally take place in double-helical DNA; for convenience only one
strand is shown.

Most common type of thymine dimer

This type of damage occurs in the DNA of cells exposed to ultraviolet radiation (as in sunlight).
A similar dimer will form between any two neighboring pyrimidine bases ( C or T residues )in
DNA.

Chemical modifications of nucleotides produce mutation

(A) Deamination of cytosine, if uncorrected, results in the substitution of one base for another when
the DNA is replicated. Deamination of cytosine produces uracil. Uracil differs from cytosine in its
base-pairing properties and preferentially base pairs with adenine. The DNA replication machinery
therefore adds an adenine when it encounters a uracil on the template strand. (B) Depurination can
lead to the loss of a nucleotide pair When the replication machinery encounters a missing purine on
the template strand it may skip to the next complete nucleotide as illustrated here ,thus producing a
nucleotide deletion in the newly synthesized strand. Many other types of DNA damage if left
uncorrected also produce mutations when the DNA is replicated.

A comparison of two major DNA repair pathways

(A) Base excision repair. This pathway
starts with a DNA glycosylase.
Here the enzyme uracil DNA
glycosylase
removes
an
accidentally deaminated cytosine
in DNAthe sugar phosphate with
the missing base is cut out by the
sequential
action
of
AP
endonuclease
and
a
phosphodiesterase. (The same
enzymes begin the repair of
depurinated sites directly). The
gap of a single nucleotide is then
filled by DNA polymerase and
DNA ligase ( apurinic sites) or by
the loss of a pyrimidine
(apyrimidinic sites)
(B) one cut is made on each side of
the lesion and an associated DNA
helicase then removes the entire
portion of the damaged strand
The excision nuclease enters and
cleaves on either side of the
damage leaving a gap of about 30
nucleotides.
The
nucleotide
excision repair machinery in both
bacteria
and
humans
can
recognize and repair many
different types of DNA damage.

The recognition of an unusual nucleotide in DNA by base

flipping.

The DNA glycosylase family of enzymes recognize specific bases in the conformation shown. Each
of these enzymes cleaves the glycosyl bond that connects a particular recognized base (yellow) to
the backbone sugar, removing it from the DNA. (A) Stick model; (B)space-filling model.

The deamination of DNA nucleotides

In each case, the oxygen atom that is

added in this reaction with water is
colored red. (A) The spontaneous
deamination products of A and G are
recognizable as unnatural when they occur
in DNA and thus are readily recognized and
repaired
(B) About 3% of the C nucleotides in
vertebrate DNAs are methylated to help in
controlling gene expression (discussed in
Chapter 7). When these 5-methyl C
nucleotides are accidentally deaminated
they form the Natural nucleotide T.
However this T will be paired with a G on
the
opposite
strand,
forming
a
mismatched base pair.

Two different ways to repair double-strand breaks

(A) Nonhomologous end-joining alters the original DNA sequence when repairing a
broken chromosome. These alterations can be either deletions (as shown) or short
Insertions. (B) Repairing double-strand breaks by homologous recombination is more
difficult to accomplish, but this type of repair restores the original DNA sequence. Lt
typically takes place after the DNA has been duplicated but before the cell has divided.

The Holliday model of DNA

crossover.
In E. coli, the recombination is
initiated by the enzyme RecBCD.
This enzyme has both helicase and
nuclease activities. The enzyme
first uses its helicase activity to
unwind DNA. When it hits the Chi
site, one of the exposed strand will
be cut by its nuclease activity

After DNA strands are cut by

RecBCD, the strand invasion is
catalyzed by RecA proteins, which
can wrap around single stranded
DNA and direct it to form the
Holliday structure.
Finally, the branch migration is
catalyzed by RuvA and RuvB. The
Holliday structure is resolved by
the protein RuvC