DNAThe "Stuff" of Life
Its replication and its
unending repair
Watson and Crick published a second paper which suggested a
hypothesis for the replication of DNA. That each strand of the DNA
molecule could act as a template for the synthesis of opposite strand
of the DNA molecule
The question arises-Is DNA replication conservative, (The old strand
is kept in tact, giving rise to two new strands), semi-conservative
(gives rise to 2 strands an old strand and a new strand) or dispersive
(new strands having both new and old strands)?
Matt Meselson and
Frank Stahl (1958)
demonstrated that
replication was semiconservative using
radioactive
nucleotides with
dividing bacteria.
The bacteria was given radioactive nucleotides so that all the DNA
was radioactive. Then once it was all radioactive, the bacteria was
given normal radioactive and then allowed to replicate once. The
DNA was then run on a agarose gel to see what banding pattern
appeared.
If replication was
conservative, it is
expected that two bands
would appear, one
radioactive and one
normal. Only one
radioactive band
appeared instead of a
two bands. This
eliminated the
conservative theory. The experiment was repeated but this time the
bacteria was able to replicate twice.
If replication was dispersive, only one band would be expected
because all the DNA would be the same weight. What appeared
though was two bands. One was radioactive and the other was not.
This eliminated the dispersive theory. DNA replication is
semiconservative theory.
The following is a
simple model for
DNA replication.
-Because DNA is such a long
molecule, replication must
occur at the same time in
many places.
This is a micrograph, three replication
bubbles are visible along the DNA cultured
Chinese hamster cells. The arrows indicate
the directions of DNA replication at the two
ends of the bubble.
There a number of proteins involved in DNA
replication.
In prokaryotic cells, the
chromosome is circular and
not linear like eukaryotic cells.
There is also only one origin
for replication which attached
to the plasma membrane.
Replication of the chromosome
occurs in both directions like
eukaryotes. Prokaryotes have
far fewer DNA base pairs than
eukaryotes. E. coli has about
4.6 million base pairs whereas
a human eukaryotic cell has 3
billion base pairs.
Notes about enzymes
-DNA polymerase can
only add to the 3' end of a
nucleotide. This means
that synthesis can only
occur from the 5'->3'
direction.
-DNA polymerase must
always have a nucleotide in front of it to hang the DNA nucleotide
on.
-Therefore an RNA primer must be laid down first and then
replaced by DNA polymerase. RNA primase does this.
-Helicases break the hydrogen bonds and gyrases relieve the stress.
-Ligases suture DNA fragments together in the replicated molecule.
-Nucleotides are always added on as triphosphates. When the
nucleotides are added then two phosphates are cleaved off making a
pyrophosphate.
1. When it is time for the DNA to replicate,
proteins attach to the places on the DNA
molecule called origins of replication. On a
prokaryotic cell there is one such site, but on a
eukaryotic cell there are hundreds to
thousands.
2. Helicases unwind the DNA strand and
gyrases will nick one of the strands to relieve
stress.
3. Single-stranded binding proteins stabilize the
unwound parental DNA strand.
4. Primase will lay down a RNA primer so that
the DNA polymerase can start base pairing
because DNA polymerase must have something to
bind to The molecule is replicated in both
directions. One side is laid down
continuously, the other side is laid down in
discontinuous fragments because and can only
grow from the 5' to the 3' direction. These
fragments are called Okazaki fragments.
5. The Okazaki fragments are joined together by
ligase.
6. The RNA nucleotides are eventually replaced
by DNA nucleotides when the DNA polymerase
runs into them.
Once the DNA has been
replicated, there is one
problem. The usual
replication machinery
provides no way to
complete the 5 ends, so
repeated rounds of
replication produce
shorter DNA molecules.
The solution to the
problem is to put on the
ends of the DNA,
repetitive sequences of
DNA. These sequences
are noncoding
sequences. These are
called telomeres.
Telomeres only provide
junk sequences so they
prevent the erosion of genes
are cells replicate.
Note- in gametes, the
shortening of telomeres
would cause serious
problems in multicellular
organisms. If chromosomes
of germ cells became
shorter in every cell cycle,
essential genes would
eventually be missing from
the gametes they produce.
An enzyme called
telomerase catalyzes the
lengthening of telomeres in
germ cells
Proofreading and Repairing
DNA
•DNA polymerases proofread
newly made DNA, replacing any
incorrect nucleotides
•In mismatch repair of DNA,
repair enzymes correct errors in
base pairing
•DNA can be damaged by
chemicals, radioactive emissions,
X-rays, UV light, and certain
molecules (in cigarette smoke for
example)
•In nucleotide excision repair, a
nuclease cuts out and replaces
damaged stretches of DNA
The
End