dnarepair2010

DNA Repair

Professor Wan Zurinah Wan Ngah

Objectives; At the end of the lectures

the students should be able to

• describe the importance of DNA repair

• Describe factors that can damage DNA

• describe the types of damage that can occur

• explain base and nucleotide excision repair processes

• explain the consequences when the damage is not repaired

DNA damage occurs

spontaneously but can increase

by various chemical, biological

and physical factors

Factors

• Damage occurs spontaneously eg. 10000 depurinations/d, spontaneous deamination of C to U (100/d)

• Other factors that can increase DNA damage:

• Biological agents eg. viruses

• Chemicals eg. mutagens, carcinogens

• Physical agents eg. Radiation, UV, X-rays

The lesion that occurs is specific

to the causative factor

Types of DNA damage/lesions

DNA Lesions Cause

Loss of base (Depurination) Acid & heat

Altered base Ionizing radiation,

alkylating agents

Wrong base Spontaneous deamination,

C to U, A to HX, 5-

Methyl C to T

Deletion-insertion Intercalating agents eg.

Acridine dyes

Thymine dimers UV

Strand breaks Chemicals, ionizing

radiation

Spontaneous changes that require DNA

repair

Spontaneous oxidative damage( red arrows)

Hydrolytic attack (blue arrows)

Uncontrolled methylation by S-Adenosyl Methionine (green arrows)

Base Deamination in DNA

Unnatural bases for DNA

Natural bases

Most frequent changes causing serious DNA

damage in cells Unnatural base

UV light increases the formation

of thymine dimers

Thymine dimers

are cyclobutyl

dimer

Carcinogens cause damage by

altering bases and DNA structure

Cytotoxic drugs also act by altering

bases and DNA structure

Example of a carcinogen from the diet

and environment

Polyaromatic hydrocarbon

group includes

benzo(a)pyrene (cigarette

smoke)

The epoxide can covalently

bind to guanine base in

DNA

Direct or complete

carcinogens

Indirect or incomplete

carcinogens

Also a carcinogen from our diet

Intercalation chemical into the DNA

double helix

Aromatic

compounds

intercalate in

between base

stacks. Results

in insertion

and addition

of new bases

Improper or incomplete DNA

repair can lead to disease

Cells invests heavily in repair

mechanisms

Importance of DNA Repair

• DNA damage DNA changes Mutation

• Mutation somatic cells Cancer

• Mutation gamete cells Genetic diseases (Inborn errors of metabolism)

• Cell invests heavily in repair enzs

• Def in repair enzs lead to diseases eg. Xeroderma Pigmentosum patients sensitive to UV, skin lesions---skin cancer etc

• Bloom Syndrome : Helicase/Ligase def, cancer risk is high

Inherited Syndromes with Defects in DNA Repair

NAME

PHENOTYPE

ENZYME OR PROCESS

AFFECTED

Xeroderma

pigmentosum (XP)

Groups A-G

Ataxia-telangiectasia

(AT)

BRCA-1 & BRCA-2

Bloom syndrome

skin cancer, cellular UV

sensitivity, neurological

abnormalities

Leukemia, lymphoma,

cellular -ray sensitivity,

genome instability

breast and ovarian

cancer

Cancer at several sites,

stunted growth,

genome instability

nucleotide excision-repair

ATM protein, a protein kinase

activated by double-strand

breaks, repair by homologous

recombination

Tumour suppressor gene,

activated by double stranded

DNA breaks

accessory DNA helicase for

replication/Ligase 1

Types of DNA Repair

• Excision repair

– Repair of thymine-thymine dimers

– Apurinic/apyrimidinic repair

– Removal of uracil

• Direct repair

– Dealkylation of G by G alkyltransferases

• Mismatch repair

• Recombination repair

Two types of excision repair are

base excision repair and

nucleotide excision repair

Excision repair

• Incision: Enz recognise type of damage. Enz

involved depends on type of damage.

– DNA N-glycosylases--deaminated base, alkylated

or oxidised bases, bases with opened rings

– Depurination--Apurinic-apyrimidinic (AP)

endonuclease;

• Excision of damaged base

• Re-synthesis of new DNA

• Nick is joined by ligase

Repair enzymes

• In Prokaryotes-DNA polymerase I (& II)

• In Eukaryotes-DNA polymerase

• Ligase

Base excision repair is for small

lesions

Nucleotide excision repair is for

large lesions such as thymine

dimers

Cyclobutyl/ thymine dimer repair

Location on DNA/gene where

mutations can occur

Mutations can occur in the:

• Coding regions or exons

• Intron/exon (splicing) junction/site

• Regulatory region

Class, Group & Type of Mutations Mutation Group Type

Substitution Synonymous Silent

Non-synonymous Missense

Nonsense

Splice site

Promoter

Deletion Multiple of 3

Not multiple of 3 Frameshift

Large deletion Partial/whole gene deletion

Class, Group & Types of Mutations:

continued

Insertion Multiple of 3

Not multiple of 3 Frameshift

Large insertion Partial/whole gene duplication

Expansion of Dynamic mutation

trinucleotide repeats

missense mutation:non-conservative substitution

a mutation results a change in an amino acid

where the new amino acids has a different property than the

old amino acid. The protein with the new primary structure

may have reduced or no activity. Or qualitative changes with

different characteristics but same biological activity

nonsense mutations:

a mutation results in a new stop translation condon

formed before the naturally occuring one.

Translation is stopped prematurely and a shortened

protein with no biological activity is made. mRNA

transcripts degraded by nonsense mediated decay

frameshift mutation: a deletion or insertion of one base results in a change in the

translational reading frame resulting in premature stop codon

downstream. mRNA maybe degraded by nonsense mediated

decay or a truncated protein is produced.

Summary

• DNA damage can be increased by biological, chemicals and physical factors.

• Lesions include loss of base, wrong base, altered base, thymine dimers, strand breaks or deletion-insertion.

• Damage is repaired by repair enzymes

• Mechanism of repair include base excision repair and nucleotide excision repair

DNA Repair

A Muslim‟s speech is not exaggerated or affected.

Adbdullah ibn Mas‟ud said:

“By Him besides Whom there is no other god, I never saw

anyone who was harsher on those who exaggerate in their

speech than the Messenger of Allah swt, and I never saw

anyone who was harsher on them after his death than Abu

Bakar, and I think that „Umar feared the most for them of

all people on earth”

Okazaki fragments have which

one of the following properties?

A They are double stranded

B They contain covalently linked RNA and DNA

C They are RNA-RNA hybrids

D They arise from the nicking of the sugar-phosphate backbone of the parental DNA chain

E They are removed by nuclease activity

Answer B

•Which one of the sequences listed below best

describes the order in which the following

enzymes participate in lagging strand DNA

synthesis in bacteria?

1 DNA polymerase I 2 5’exonuclease

3 DNA polymerase III 4 DNA ligase

5 RNA polymerase

A 5,1,3,2,4

B 3,2,1,5,4

C 5,3,4,2,1

D 5,3,2,1,4

E 3,2,5,1,4

ANSWER D

Formation of thymine dimers

Questions

• List the factors that can cause damage to

DNA

• How can cigarette smoke cause cancer?

• List 6 types of lesions that can occur to DNA.

• How can dietary contaminants cause

damage to DNA?