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chapter 24
DNA Replication, Repair, and Mutagenesis
24.1
DNA Replication
Problems of Replication
The replication process requires that each double-helical
molecule of DNA produce two identical molecules of
DNA. This means that wherever a G-C or A-T base pair
occurs in the parental molecule, the identical base pair
must occur in the progeny molecules. However, many fac-
tors interfere with accurate replication of DNA. If an A
should pair with C or G with T as a result of tautomeriza-
tion (Chapter 23), a
p o in t m u ta tio n
(a change in one base
pair) will result. Occasionally, a segment of DNA will be
replicated more than once (
d u p lic a tio n
) or a segment may
fail to be replicated
(deletion ).
These and other aberra-
tions in DNA replication do occur, but the mechanism of
replication has evolved to minimize such mistakes.
Semiconservative Replication
The information in each strand of the double helix serves as
the template for the construction of a new double-helical
DNA molecule; this is called
sem ico n serva tive rep lica -
tion
since one old strand of DNA is paired with one new
strand to produce the daughter DNA molecule. All DNA
molecules in all organisms replicate semiconservatively.
This basic fact was first proved by Matthew Meselson
and Frank Stahl who labeled DNA in
E. coli
by growing
cells for several generations in medium containing a heavy
isotope of nitrogen (
1 5
N). Cells were then transferred to
medium containing 14N and continued to grow exponen-
tially. After one generation of growth in l4N medium, the
DNA was extracted from a sample of cells and analyzed by
cesium chloride density centrifugation. All DNA was of
intermediate (hybrid) density; one strand contained 15N
and the newly synthesized strand contained
1 4
N. After
two generations of growth in l4N medium, 50% of the
DNA contained only I4N (light DNA) and 50% is hybrid.
This important experiment confirmed a prediction of the
Watson-Crick model for DNA. Subsequent experiments
also showed that human DNA undergoes semiconserva-
tive replication.
Origin and Direction of DNA Replication
Another unifying principle of DNA replication is that each
molecule of DNA has one or more specific
orig in s o f re p li-
cation
where DNA synthesis begins. In bacteria that carry
their genetic information in a circular DNA molecule, only
one origin of replication exists. Plasmid DNA also con-
tains a unique origin of replication. Origins of replication
can be moved from one location in DNA to another, or
TABLE 24-1
C o m p a r is o n o f P a r a m e te r s o f D N A R e p lic a tio n in
E . c o li a n d H u m a n C ells
DNA
E. co li
Human
Amount of DNA
~3.9x 10
6
bp
~3x
1 0 9
bp
Rate of replication at
-850 bases/sec
-6-90 bases/sec
each replication fork
Number of origins of
one
1 0 3
to
1 0 4
replication
Time for cell division
-30 min
-24 hours
from one DNA molecule to another just as genes can be
moved by genetic engineering techniques.
In both bacterial and plasmid DNA, replication is ini-
tiated at a unique origin and proceeds in both directions
along the DNA molecule to a
term in a tio n site
that is lo-
cated approximately 180 degrees from the origin. Thus,
replication of DNA is, in most cases,
b id irectio n a l rep li-
cation .
The ability of DNA to replicate from many origins
and in both directions means that cells can replicate all of
their DNA in a fairly short period of time. The shortest
generation time for the bacterium
E. coli
is approximately
30 minutes in rich medium in the laboratory whereas the
shortest division time for a human cell is approximately
24 hours (Table 24-1).
Unwinding a double helix during replication presents a
serious mechanical problem. Either the two daughter DNA
molecules at the
rep lica tio n fo r k
(the Y-shaped fork) must
rotate around one another, or the unreplicated segment of
DNA must rotate (Figure 24-1). This necessity for DNA
rotation during replication creates topological problems
for covalently closed circular DNA in bacteria and for the
enormously long condensed and folded DNA in human
chromosomes. The unwinding of strands of the DNA dou-
ble helix is accomplished by a special group of enzymes
called
h elicases;
positive and negative coiling of DNA to
maintain the topology necessary for replication is the func-
tion of another group of enzymes called
to poisom erases
(discussed in a later section).
Replicons
Much of the basic understanding of DNA replication was
gleaned from studying the replication of
p la sm id s
(Chap-
ter 23): small, circular, extrachromosomal DNA elements
in bacteria. Some plasmids, such as the F (fertility) factor
in
E. coli,
have an origin of replication but replicate only
unidirectionally from that single origin by a rolling-circle
mechanism of replication (Figure 24-2). If an F factor
