Page 68 - Genetics_From_Genes_to_Genomes_6th_FULL_Part2

7.2 Molecular Mechanisms That Alter DNA Sequence 227

found in RNA but not in DNA. Because U pairs with A Figure 7.9 DNA polymerase’s proofreading function. If
rather than G, deamination followed by replication may DNA polymerase mistakenly adds an incorrect nucleotide at the
alter a C–G base pair to a T–A pair in future generations 3′ end of the strand it is synthesizing, the enzyme’s 3′-to-5′
of DNA molecules (Fig. 7.8b); such a C–G to T–A exonuclease activity removes this nucleotide, giving the enzyme a
second chance to add the correct nucleotide.
change is a transition mutation.
Other assaults include naturally occurring radiation 3' – 5' exonuclease Wrong base
cuts here
such as cosmic rays and X-rays, which break the sugar- added
phosphate backbone (Fig. 7.8c); ultraviolet light, which 5' A Template
strand
causes adjacent thymine residues to become chemically CCC AA T GGT
linked into thymine dimers (Fig. 7.8d); and oxidative
damage to any of the four bases (Fig. 7.8e). If not repaired 3' GGG TT A CC A GA AC GT A T 5'
before DNA replication, all of these changes alter the infor-
mation content of the DNA molecule permanently. DNA polymerase
Wrong base
excised
5'
Mistakes in DNA Replication Also CCC AA T GGT
Cause Spontaneous Mutations GGG TT A CC AG AA CG T A T
If the cellular machinery for some reason incorporates an 3' 5'
incorrect base during replication, for instance, a C oppo-
site an A instead of the expected T, then during the next DNA polymerase
can now add the
replication cycle, one of the daughter DNAs will have the correct base
normal A–T base pair, while the other will have a mutant 5'
G–C. Careful measurements of the fidelity of replication CCC AA T GGT C T
in vivo, in both bacteria and human cells, show that such GGG TT A CC AG AA CG T A T
errors are exceedingly rare, occurring less than once in 3' 5'
9
every 10 base pairs. That rate is equivalent to typing this
entire book 1000 times while making only one typing
error. Considering the complexities of helix unwinding,
base pairing, and polymerization, this level of accuracy is error rate collectively another 10-fold, bringing it to
amazing. How do cells avoid most DNA replication within about 100-fold of the fidelity attained by the
errors, and what kinds of mistakes occur nonetheless cell. The 100-fold higher accuracy of the cell depends
when DNA is copied? on a backup system called methyl-directed mismatch
repair that notices and corrects residual errors in the
newly replicated DNA. We present the details of this
The proofreading function of DNA polymerase repair system later in the chapter when we describe the
The replication machinery minimizes errors through suc- various ways in which cells attempt to correct muta-
cessive stages of correction. In the test tube, DNA poly- tions once they occur.
merases replicate DNA with an error rate of about one
6
mistake in every 10 bases copied. This rate is about
1000-fold worse than that achieved by the cell. Even so, it Base tautomerization
is impressively low and is attained only because poly- One reason why DNA polymerase may make mistakes
merase molecules provide, along with their polymeriza- is the tautomerization of bases. Each of the four bases
tion function, a proofreading/editing function in the form has two tautomers, similar chemical forms that inter-
of a nuclease that becomes active whenever the poly- convert continually. The equilibrium between the tauto-
merase makes a mistake. This nuclease portion of the mers is such that each base is almost always in the form
polymerase molecule, called the 3′-to-5′ exonuclease, in which A pairs with T and G pairs with C. However, if
recognizes a mispaired base and excises it, allowing the by chance a base in the template strand is in its rare
polymerase to copy the nucleotide correctly on the next tautomeric form when DNA polymerase arrives, the
try (Fig. 7.9). Without its nuclease portion, DNA poly- wrong base will be incorporated into the newly synthe-
merase would have an error rate of one mistake in every sized chain because the rare tautomers pair differently
4
10 bases copied, so this editing function improves the than do the normal forms (Fig. 7.10a). If the misincor-
fidelity of replication 100-fold. porated nucleotide is not corrected by mismatch repair
DNA polymerase in vivo is part of a replication sys- before the next round of replication, a point mutation
tem including many other proteins that improve on the results (Fig. 7.10b).