Chapter 9 The mutability and repair of DNA

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Chapter 9 The mutability and repair of DNA

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• 200431060049

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Outline

• replication errors and their repair
• DNA damage
• repair DNA damage

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Introduction

• Organisms can survive only if their DNA is
  replicated faithfully and is protected from
  chemical and physical damage that would change
  its coding properties.The limits of accurate
  replication and repair of damage are revealed by
  the nature mutation rate.
• Thus,an average nucleotide is likely to be
  changed by mistake only about once every 109
  times it is replicated.

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Two important sources of mutation

• inaccuracy in DNA replication
• chemical damage to the genetic material

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Errors in replication and damage have two
consequences

• 1.Permanent changes to the DNA (mutations),
• can alter the coding sequence of a gene or its
  regulatory sequences.
• 2.Some chemical alterations to the DNA prevent
  its use as a template for replication and
  transcription.

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• Since the mutation is so important for all the
  living things,now we will consider errors that
  occur during replication and how they are
  repaired. We will see that multiple overlapping
  systems enable the cell to cope with a wide range
  of insults to DNA,underscoring the investment
  that living organism make in the preservation of
  the genetic material.

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Part ? Replication errors and their repair
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The nature Mutations

• Mutations include almost every conceivable change
  in DNA sequence. The simplest mutations are
  switches of one base for anther. There are two
  kindsThansitions and
• Transversions.

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• Other kinds of mutation cause more drastic
  changes in DNA,such as extensive insertions and
  deletions and gross rearrangements of chromosome
  structure. Such change may be caused by
  transposon.
• One kind of sequence that is particularly prone
  to mutation merits special comment because of its
  importance in human genetics and disease. Like
  DNA microsatellites.

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Some replication errors escape proofreading

• As we have seen,the 3-5 exonuclease component
  of the replisome,which removes wrongly
  incorporated nucletides. The proofreading
  exonuclease is not however,foolproof. Some
  misincorporated nucleotides escape detection and
  become a mismatch between newly synthesized srand
  and the template srand.

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• A mutation can be permanently incorporated by
  replication. In the second round the mutation
  becomes permanently incorporated in the DNA
  sequence

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Mismatch repair removes errors that escape
proofreading

• Mismatch repair system can detect mismatches
  and repair them. There are two challenges1,it
  must scan the genome for mismatches,it will
  rapidly find and repair .
• 2.the system must correct the mismatches
  accurately,it must replace the misincorporated
  nucleotide in the newly synthesized stand not the
  parental strand

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• In E.coli,mismatches are detected by a dimer of
  the mismatch repair protein MutS. MutS scans the
  DNA,recognizing mismatches from the distortion
  they cause in the DNA backbone. MutS embraces the
  mismatch-containing DNA, inclosing a pronounced
  kink in the DNA and a conformational change in
  itself.A key to the specificity of MutS is that
  DNA containing a mismatch is much more readily
  distorted than properly base-paired DNA.

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E.Coli tags the parental strand by transient
hemimethylation

• E.Coli enzyme Dam methylase methylates a residues
  on both strands of the sequences 5-GATC-3.when
  a replication fork passes through DNA that is
  methylated at GATC sites on both strands,the
  resulting daughter DNA duplexes will be
  hemimethylation.

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Dam methylation at replication fork(a)replication
generates hemimethylated DNA in E.coli(b)MutH
makes incision in unmethylated daughter strand
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• Eukaryotic cells also repair mismatches and do so
  using homologs to MutS and MutL. Indeed,
  eukaryotic have multiple MutS-like proteins with
  different specificities.

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• Part ? DNA damage

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DNA undergoes damage spontaneously from
hydrolysis and deamination

• Some damage is caused by environmental factors
  such as radiation and mutagens.
• The most frequent and important kind of
  hydrolytic damage is deamination of base
  cytosine.
• The hazard of having deamination generate a
  naturally occurring base is illustrated by the
  problem caused by presence of 5-methylcytosine.

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DNA is damaged by alkylation oxidation and
radiation

• DNA is vulnerable to damage from alkylation
  oxidation and radiation .
• In alkylation,methyl or ethyl groups are
  transferred to reactive sites on the bases and to
  phosphates in the DNA backbone.
• DNA is also subject to attack from reactive
  oxygen species

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Thymine dimer.uv includes the formation of a
cyclobutane ring between adjacent thymines.
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• Another type of damage to bases is caused by
  ultraviolet light.Reaction with a wavelength of
  about 260 nm is strongly absorbed by the bases
• Gamma radiation and X-rays are particularly
  hazardous because they cause double-strand breaks
  in the DNA.

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Mutations are Also Caused by Base Analogs and
Intercalating Agents

• Mutations are also caused by components that
  substitute for normal bases or slip between the
  bases to cause errors in replication.
• Base analogs are structurally similar to proper
  bases but differ in ways that make them
  treacherous to the cell .

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Base analogue of thymine,5-bromouracil,can
mispair with guanine.
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• TOPIC ? Repair of DNA damage

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• In this section, we consider the systems that
  repair damage to DNA. In the most direct of these
  systems ,a repair enzyme simply reverses the
  damage. One more elaborate step involves excision
  repair systems, in which the damaged nucleotide
  is not repaired but removed from the DNA.
• In excision repair systems, the other, undamaged,
  strand serves as a template

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• for reincorporation of the correct nucleotide by
  DNA polymerase.
• More elaborate is recombinational repair,which is
  employed when both strands are damaged as when
  the DNA is broken. In such situations, one strand
  cannot serve as a template for the repair of the
  other. Hence in recombinational repair ,sequence
  information is retrieved from a second undamaged
  copy of the chromosome

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Direct Reversal of DNA Damage

• An example of repair by simple reversal of damage
  is photoreactivation.
• Photareactivation directly reverses the formation
  of pyrimidine dimers that result from ultraviolet
  irradiation.

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Photoreactivation
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• Another example of direct reversal is the removal
  of the methyl group from the methylated
  O6-methylguanine .

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Base Excision repair enzymes remove damaged
bases by a base-flipping mechanism

• The most prevalent way in which DNA is cleansed
  of damaged bases is by repair systems that remove
  and replace the altered bases.
• The base excision repair, an enzyme called a
  glycosylase recognizes and removes the damaged
  base by hydrolyzing the glycodic bond.the
  resulting abasic sugar is removed from the DNA
  backbone in a further endonucleolytic step.

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Base excision pathwaythe uracil glycosylase
reaction
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• endonucleolytid cleavage also removes
  apurinic and apyrimidinic sugars that arise by
  spontaneous hydrolysis. After the damaged
  nucleotide has been entirely removed from the
  backbone, a repair DNA polymerase and DNA ligase
  restore an intact strand using the undamaged
  strand as a template.

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• Dleansing the genome of damaged bases is a
  formidable problem because each base is buried in
  the DNA helix. Evidence indicates that these
  enzymes diffuse laterally along the minor groove
  of the DNA until a specific kind of lesion is
  detected.
• X-ray crystallographic studies reveal that the
  damaged base is flipper out so that it projects
  away from the double helix,where it sits in the
  specificity pocket of the glycosylase.

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Nucleotide excision repair enzymes cleave damaged
DNA on either side of the lesion

• Unlike base excision repair ,the nucleotide
  excision repair enzymes don't recognize any
  particular lesion. rather ,this system works by
  recognizing distortions to the shape of the
  double helix

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Nucleotide excision repair pathway
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• Not only the nucleotide excision repair capable
  of mending damage throughout the genome, but it
  is also capable of rescuing RNA polymerase, the
  progression of which has been arrested by the
  presence of alesion in the transcribed strand of
  a gene. this phenomenon ,known as
  transcription-coupled repair, involves
  recruitment to the stalled RNA polymerase of
  nucleotide excision repair proteins .

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Recombination repairs DNA breaks by retrieving
sequence information from undamaged DNA

• Excision repair uses the undamaged DNA strand as
  a template to a replace a damaged segment of DNA
  on the other strand.
• This is accomplished by the double-strand
  break(DSB) repair pathway, which retrieves
  sequence information from the sister chromosome.
• DNA recombination also helps to repair errors in
  DNA replication.

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