Transcript Mutations

Mutations
• Mutations are inheritable changes in
the DNA
– “Failure to faithfully store genetic
information”
• Changes can be to chromosomes or
genes
– Current focus: changes to DNA
sequences.
• This means an alteration in a basepair or
• in the order of the basepairs.
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Types of mutations-1
• Mutations can be classified in many, many ways
– Some ways mutually exclusive, some not.
• Spontaneous vs. Induced
– Spontaneous happens naturally
• Enzymatic errors, especially in copying
• Various chemical reactions
– Induced mutations: specifically caused, as by researcher
• Treatment with various chemicals, radiation
• Gametic (germ line) vs. somatic
– Gametic mutations can be passed on to next generation
– Somatic only affects individual (in metazoans)
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Types of mutations-2
• Morphological
– Change in physical structure, readily observed
• Nutritional/biochemical
– Mutated enzyme results in phenotypic change
– Bacterial auxotrophs; sickle cell anemia
• Behavioral mutations
• Regulatory mutations
– Affect control of gene expression rather than protein
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Types of mutations-3
• Lethal mutations: not easily studied unless:
• Conditional mutations: expressed depending on
environmental conditions, especially temperature.
– a way to study lethal mutations: permissive and
restrictive temperatures; esp. useful with bacteria
– temp sensitive mutations occur
naturally, continued in offspring:
Siamese cats, Himalayan rabbits
www.tcainc.org/photos/ farpoint/saavik1.jpg
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Types of mutations-4
Classification of mutations by FUNCTION
– Loss of function: knockout or null.
– Hypomorphic: lowered expression, “leaky”
– Hypermorphic: greater activity or more visible trait
• typically regulatory mutation, results in increased
expression
– Gain of function: e.g. new enzymatic activity
• a factor in evolution;
– Dominant negative: bad apple spoils the bunch e.g. bad
protein in multicomponent enzyme
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Detection of mutations
• Bacteria and fungi
– Prototrophs and auxotrophs: microbe no longer able to
synthesize or breakdown particular nutrient.
– Change in behavior, e.g. motility
• Various methods in plants and animals
• Humans (“not suitable experimental organisms”)
– Reliance on pedigrees
– Possible to determine sex linkage, dominance
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Mutations are rare (but not equally so)
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• Mutation rate depends on species and on gene
• Hot spot: a location in DNA where mutations occur
significantly more often than the usual 1/ 106.
– Monotonous run of single nucleotide or tandem repeats:
GGGGGGGGG or ATGGATGGATGG
– Methylated cytosines
• methylation is added a CH3 group to something
• Cytosines are methylated to help indicate which DNA
strand is older (helps with DNA repair).
• Problem occurs when a cytosine is chemically
damaged by deamination. (more later)
Mutations
• Our example:
– information, 3 letters at a time, read consecutively
• Point mutations:
• Frameshift mutations: Insertion
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more Mutations
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• Frameshift: deletion
• Transposon mutagensis: transposons are segments of
DNA that can jump into another spot in the DNA;
they have information.
More types of mutations
• Switch between A & G, or C & T: transition
• Switch between purine and pyrimidine: transversion
• Silent: 3rd position of codon usually means same amino
acid, so change here has no effect.
• Missense: typically a single nucleotide change, causes
change in amino acid and noticeable effect.
• Nonsense: change amino acid codon to STOP codon
• Additions, deletions, and “stuttering”
– Stuttering: repeated sequences sometimes copied
incorrectly; enzyme gets confused?
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Mutagens: that which causes mutations
• Base analogs: e.g. 5-bromouracil. In equilibrium between
keto and enol forms
– In keto form, looks like T
– In enol form looks like C
– Used one way, but when copied, mispairing can occur.
• Modifying agents: chemically change bases
– HNO2 nitrous acid: deaminates (amino to keto)
• See upcoming slide: deamination
– Alkylating agents (ethylmethane sulfonate): add methyl
or ethyl group to bases (-CH3 or CH2CH3), cause
mispairing during synthesis
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Loss of a purine, a natural process
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Can lead to an
incorrect base being
added; a mutation.
saturn.roswellpark.org/.../ AP_site_generation.gif
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Pyrimidines and deamination
Deamination:
Loss of an
amine group,
replacement w/
a keto group.
Deamination of cytosine makes uracil; recognized as wrong
and repaired. Deamination of 5-methyl cytosine produces
thymine, which is normal; results in a transition mutation.
Frameshift mutations
• Cause misalignment during DNA replication; caused by
intercalating agents such as ethidium bromide or acridine
orange
http://www.photobiology.com/photoiupac2000/pierard/intintercal.jpg
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Radiation
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• UV light at 265 nm
– causes thymine dimers;
covalent connections between
adjacent thymines. Hurried
repair makes mistakes.
• Ionizing radiation
– short wavelength, high energy
radiation, e.g. x-rays, gamma
radiation.
– Causes ss, ds breaks in DNA.
http://academic.brooklyn.cuny.edu/
biology/bio4fv/page/molecular%
20biology/mutation-prym-dimers.jpeg
Ionizing Radiation
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• Major damage is from free radicals
– Most abundant substance in cell is water; radiation
produces radicals that attack DNA, causing breaks.
• The effects of radiation are a matter of considerable
scientific and political controversy.
– Effects of high levels of radiation are well understood,
but effects of low levels are very difficult to study.
– Brief soapbox: after Chernobyl tragedy, people vacated
many square miles around damaged reactor. Now,
endangered animals making a comeback despite
radiation.
Repair of DNA damage
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• Despite the constant bombardment of DNA with radiation
and chemicals, cells possess repair mechanisms.
• Repair systems exist for
– UV light damage
– Chemical changes to bases
– Loss of bases
– Incorrect copying
– Ss and ds breaks in DNA
http://earthobservatory.nasa.gov/Library/UVB/Images/dna_mutation.gif
Repair of Thymine dimers
Photoreactivation: Enzyme
uses a photon of blue light to
separate thymines from each
other. (When using UV as a
mutagen, put cells in dark
afterwards!) (in E. coli)
Excision Repair: DNA repair
enzymes recognize a distorted
DNA helix (such as caused by
thymine dimers). The entire
local section of DNA is
removed and replaced. In all prokaryotes & eukaryotes.
http://www-personal.ksu.edu/~bethmont/excisio3.gif
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Repair of chemical changes
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• Deamination of cytosine
– as shown previously, converts cytosine to uracil
– the enzyme uracil glycosylase cuts off uracil, leaving
deoxyribose as part of backbone, creating an “AP” site
• AP = apurinic or apyrmidinic, meaning purine etc.
NOT there.
– AP repair, mechanism that specifically fixes such places.
Creation of an AP site
saturn.roswellpark.org/.../ AP_site_generation.gif
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AP repair- continued
Activity of uracil glycosylase or
spontaneous loss of base from DNA
can create an AP site.
An endonuclease cuts out the
remaining sugar-phosphate and
replaces it with
a complete nucleotide.
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Bulky excision repair
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• Like in repair of UV-induced damage, cells sense
bulges, kinks, or similar damage to DNA
• Chunk of DNA containing the damaged area is
excised, replaced by DNA polymerase I enzyme (or
equivalent)
– 13 bases removed in bacteria
– Eukaryotes (always more elaborate) take out 28
nucleotides
– In humans, failure in this repair system causes disease
xeroderma pigmentosum with increased risk of skin
cancer.
Mismatch repair
If Proofreading misses:
Other enzymes recognize
that the wrong base pair is
in place, cuts out
incorrect one and replaces
it. Which one is incorrect?
Presumably the newest
one = the one on the DNA
chain with the least
amount of methyl
cytosines.
cmgm.stanford.edu/.../DNA%20Repair%20-%20Doug/
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SOS Repair
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• Especially in bacteria, when damage to DNA is
severe, an emergency system goes into effect where
damage is repaired rapidly, but sloppily. Introduces
many mutations, some possibly fatal, but DNA
damage would surely be fatal otherwise.