Chapter 9 The Mutability and Repair of DNA.
Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author.While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server.
Mutations include almost every conceivable change in DNA sequence.
point mutation is the mutation that alter a single nucleotide .
a . Transitions (pyrimidine to pyrimidine, purine to purine)
b .Transversions (pyrimidine to purine, purine to pyrimidine
Gross rearrangement of chromosome
“hotspots”: the sites on the chromosome where the mutations arise at high frequency. (eg. DNA microsatellite ,Mutation-prone sequence in human genome are repeats of simple di-, tri- or tetranucleotide sequences, these sequences (1) are important in human genetics and disease, (2) hard to be copied accurately and highly polymorphic in the population )
2.Three important sources for mutation (unavoidable)
3.Two consequences of mutation unrepaired(1)Permanent changes to DNA which can alter the coding sequence of a gene or its regulatory sequence.(2)Some chemical alterations to DNA prevent its use as a template for replication and transcription.So repair of DNA is important to the organisms.OUTLINE
As we have learned ,the proofreading mechanism ( the 3’→5’ exonuclease component) improves the fidelity of DNA replication, but it is not foolproof.
So there are a total of 12 possible mismatches during the DNA replication (T:T,T:G,T:C, and so forth)
The misincorporated nucleotide needs to be detected and replaced, otherwise it will cause mutation.
Now let us discuss the process of repair in detail.
Firstly,E. coli’s repair system
1.How to scan the mismatches and remove errors?
2.How to distinguish the mismatched strand and the parental strand?
MutS scans the DNA, recognizing the mismatch from the distortion they cause in the DNA backbone.
MutS is a dimer of the mismatch repair protein.
MutS embraces the mismatch-containing DNA, inducing a pronounced kink in the DNA and a conformational change in MutS itself.
MutS-mismatch-containing DNA complex recruits MutL, MutL activates MutH, an enzyme causing an incision or nick on one strand near the site of the mismatch.
MutL is a second protein component of the repair system.
MutH is an enzyme causing an incision or nick on one strand
The helicase (UrvD) unwinds the DNA starting from the incision and moving in the direction of the site of the mismatch , and the exonucleases progressively digests the displaced single strand , extending to and beyond the site of the mismatched nucleotid.
This action produces a single-stranded gap , which is then filled in by DNA polymerase Ⅲ and sealed with DNA ligase.
The answer is that E. coli tags the
parental strand by transient
hemimethylation as we now
The GATC sequence is widely distributed along the entire genome ,and all of these sites are methylated by the Dam methylase. So before the newly strand is methylated by the Dam methylase after the DNA replication ,the resulting daughter DNA duplexes will be hemimethylated , thus the newly strand is marked (it lacks a methyl group) and hence can be recognized as the strand for repair .
Different exonucleases are used to remove ssDNA between the nick created by MutH and the mismatch dependoing on whether MutH cuts the DNA in the 5’ or 3’ side of the misoncorporated nucleotide .
As we see in chapter 8, takes place discontinuously wiyh the formation of Okazaki fragments that are joined to previously synthesized DNA by DNA ligase. Prior to the ligation step ,the Okazaki fragment is separated from previously synthesized DNA by a nick created, which can be though of as being equivalent to the nick created in E.coli by MuntH on the newly synthesized strand.OUTLINE
DNA undergoes damage spontaneously from hydrolysis and deamination.
DNA damaged by alkylation , oxidation and radiation.
Mutations are also caused by base analogs and intercalating agents.
The most frequent and important kind of hydrolytic damage is deamination of the base cytosine, (just show in the picture on the left)
Notice that , in contrast to the replication errors ,all of these hydrolytic reactions result in alterations to the DNA that are unnatural.
In alkylation ,methyl
or ethyl groups are transferred to reactive
sites on the baese and to phosphates in the DNA backcone.
DNA is also subject to attack from reactive oxygen species .(eg. O2-, H2O2, and OH•)
ultraviolet light ,which produce the photochemical fusion of two pyrimidines that occupy adjacent positions on the same polynucleotide chain. (eg. thymine dimer) These linked bases are incapable of base-pairing and cause the DNA polymerase to stop during replication.
gamma radiation and X-ray, which cause double-strand breaks in the DNA ,which are different to repair.
Similar enough to the normal bases to be processed by cells and incorporated into DNA during replication.
But they base pair differently, leading to mistake during replication.
They are flat molecules containing several polycyclic rings that rings that bind to the equally flat purine or pyrimidine bases of DNA, just as the bases bind or stack with each other in the double helix. OUTLINE
DNA repair system
removes the methyl group
from the methylated
O6-methylguanine . The methyl group is transferred to the protein itself, inactivating the protein. (very costly)
A dedicated glycosylase which recognizes the oxoG:A base pairs recognizes an A opposite an oxoG as a mutation and removed the undamaged but incorrect base.
(b) UvrA leaves the
UvrB melts DNA
(c)UvrC forms a
complex with UvrB
and creates nicks to
5’ side of the lesion
and to the 3’ side of
the lesion .
(d)DNA helicase UvrD
releases the single
from the duplex ,and
DNA pol 1and ligase
repair and seal the
that has been
arrested by the
lesions in the DNA
chromosome is present in the
cell , the DSB-repair (double
strand break system) pathway
can operate. the DSB-repair
retrieves sequence information
from the sister chromosome. (Details are in chapter 10)
known as NHEJ
(nonhomologous end joining) .
NHEJ does not involve homologous recombination, instead, the two ends of broken DNA are directly joined to each other by misalignment between single strand protruding from the broken ends.
rate, translesion synthesis can be
considered a system of last resort.
1.How does the translesion polymeras recognize a stalled replication fork ?
2.How does the translesion enzyme replace the normal replication polymerase in the DNA replication complex?
3.Once DNA synthesis is extended across the lesion ,how does tge normal replication polymerase switch back to and replace the translesion enzyme at the replication fork? OUTLINE
1.How is the DNA mended
rapidly enough to prevent
errors from becoming set in the
genetic material as mutation?
2.How does the cell distinguish the
parental strand from the daughter
strand in repairing replication errors?
3.How does the cell restore
the proper DNA sequence
when the original sequence
can no longer be read?
4.How does the cell deal with
lesions that block replication?
we have discussed errors that are generated during replication , lesions that arise from spontaneous damage to DNA , and the damage that is wrought by chemical agents and radiation.
In my opinion , in each case we should consider what cause the alteration to the genetic material , how the alteration to the genetic material is detected and how it is properly repaired. Then if we understand these problems clearly, we must make it!