Biology Reference
In-Depth Information
the RNA-DNA primers initially synthesized by the polymerase
α
-primase com-
plex. A DNA polymerase fills the gaps between the Okazaki fragments after the
primers are removed (
Sutton and Walker 2001, Hubscher et al. 2002
).
Proteins (called sliding-clamp proteins and clamp-loading proteins) act at the
eukaryote replication fork to load the polymerase onto the primer and maintain
its stable association with the template. The
clamp-loading proteins
(called rep-
lication factor C) recognize and bind DNA at the junction between the primer
and template. The
sliding-clamp proteins
(proliferating cell nuclear antigen) in
eukaryotes bind adjacent to the clamp-loading proteins, forming a ring around
the template DNA. The clamp proteins then load the DNA polymerase onto the
DNA at the primer-template junction.
The ring formed by the sliding clamp maintains the association of the poly-
merase with its template as replication progresses, allowing the uninterrupted
synthesis of long DNA molecules. Helicases unwind the template DNA ahead
of the replication fork. Single-stranded DNA-binding proteins (eukaryotic rep-
lication factor A) then stabilize the unwound template DNA so that the single-
stranded DNA can be replicated. The enzymes involved in DNA replication, in
combination with their accessory proteins, synthesize both leading and lagging
strands of DNA simultaneously at the replication fork. The idea that DNA poly-
merases track like locomotives along the DNA template during DNA replication
is pervasive and is probably based on the misperception that the polymerase is
smaller than the DNA (
Cook 1999
). We now know that the DNA polymerase-
protein complexes involved in DNA replication can be much larger than the DNA
template.
An alternative model to the “movement” of polymerase along the DNA tem-
plate has been proposed in which the fixed polymerase complexes “reel in their
DNA templates” as they extrude newly made DNA in replication “foci” or repli-
cation factories within the cell. This “fixed” model assumes that the DNA poly-
merase complex is fixed and that the DNA rotates around it. This solution is a
simple solution to the potential problem of untangling DNA strands that twine
around each other if the DNA polymerase moves (
Cook 1999
).
Some DNA polymerases in eukaryotes have 3
′
to 5
′
exonuclease activity in
addition to their polymerase activity, meaning that they can excise a misincor-
porated nucleotide by (proofreading) during DNA replication. DNA mismatch
correction further minimizes replication errors by a survey of newly synthesized
DNA strands. Furthermore, accessory factors such as DNA helicases apparently
improve accuracy during DNA elongation, possibly due to resolution of stalled
replication forks. Despite all these precautions, occasional misincorporated
nucleotides or deletions, or insertions may remain, resulting in mutations.
