DNA replication and repair for medical students

1. Dr.S.Sethupathy,M.D.,Ph.D,
Professor & Head of Biochemistry
RMMC
AU

2. Deoxyribo Nucleic Acid (DNA)
 Deoxyribose sugar
 Nitrogenous bases
 Purine bases- Adenine and Guanine
 Pyrimidine bases- Thymine and cytosine
 Phosphoric acid
 Base + sugar = nucleoside
 Base + sugar + phosphoric acid = Nucleotide

3. DNA

4. DNA structure

5. Deoxy nucleosides and nucleotides
Deoxy adenosine, deoxy guanosine,
dexoy cytidine, deoxy thymidine
Deoxy adenylate, deoxy guanylate,
dexoy cytidylate, thymidylate
(dAMP, dGMP, dCMP, TMP)

6. Double stranded
Polarity
Antiparallel
Equal number of adenine and thymine
bases
Equal number of guanie and cytosine
bases
Chargaff’s base pairing rule
A .. T and G … C

7. Sugars

8. RNA vs DNA

9. Difference between DNA & RNA

10. RNA

11. Transfer RNA

12. messengerRNA

13. Dr.S.Sethupathy, M.D.,Ph.D.,
Professor & HOD of Biochemistry,
Rajah Muthiah Medical College,
Annamalai University.

14. DNA replication
 Copying of DNA – DNA from DNA
 Only once
 Semi conservative
 Each strand acts as template
 Polarity is maintained.
 Reading 3’ to 5’
 Synthesis 5’ to 3’

16. Steps of replication
 Identification of Origin
 Unwinding of DNA
 Replication fork and RNA primer synthesis
 Synthesis and elongation of DNA - Polymerization
 Replication bubbles and ligation of gaps
 Chromatin assembly formation

17. Origin of replication
E.coli – Dna A protein finds
Ori C
Yeast- Origin Recognition
complex (ORC)
Eukaryotes - DNA
unwinding element (DUE)

19. Replication fork
 As DNA synthesis continues, there is a replication
fork with two prongs.
 Eukaryotes have replication at multiple origins
 DNA helicase unwinds DNA
 Helicases - separate strands of a DNA double helix
 Also remove nucleic acid-associated proteins.

21. Single-strand binding proteins bind to
the DNA (SSBPs)
(Gyrase)Topisomerases I and II –
removes supercoils by nicking and
sealing.
RNA primer
11- 60 nts length
Dna G synthesize – RNA polymerase
activity
Leading strand – one
Lagging strand – many

22. DNA polymerase
Reading 3’ to 5’
Synthesis 5’ to 3’
Leading strand (forward) –
continuous
The lagging strand is synthesized
in short segments as Okazaki
fragments.

23. DNA base pairing

24. Termination
 In Eukaryotes DNA replication is unable to reach the
very end of the chromosomes.
 The telomere of the daughter DNA strand shortens.
 So somatic cells can only divide a certain number of
times.
 In germ cell line, telomerase enzyme extends the
repetitive sequences of the telomere region to prevent
degradation.
 Telomerase if active in somatic cells, leads to cancer.

26. DNA ligase
DNA ligases close nicks in the
phosphodiester backbone of DNA.
 DNA ligases are essential for
joining Okazaki fragments .
Involved in DNA repair process.

27. Reverse transcriptase
 In retroviruses, the RNA is the template.
 Reverse transcriptase – DNA from RNA
 DNA-RNA hybrid is synthesized
 RNAase H degrades the RNA
 Then dsDNA is synthesized from DNA
template.
 Retroviruses (e.g., HIV), and RT inhibitors
are widely used as antiretroviral drugs.

28. Cell cycle

30. Clinical applications
 A disregulation of the cell cycle lead to tumor
formation.
 Cell cycle inhibitors, RB, p53 etc. on mutation, form a
tumor.
 G1 is the most variable phase of the cell cycle.

31. DNA repair
 DNA damage needs repair.
 Ineffective repair lead to three possible states:
1. an irreversible state of dormancy, known as senescence
2. cell suicide, also known as apoptosis or programmed
cell death
3. unregulated cell division, which can lead to the
formation of a tumor that is cancerous

32. Types of DNA damage
1. Endogenous damage - attack by reactive oxygen
species produced from metabolism.
2. Exogenous damage by external agents
ultraviolet , X-rays and gamma rays
Thermal disruption, toxins, mutagenic chemicals,
aromatic compounds that intercalate DNA
viruses.

34. 1.Single base alteration
2.Two base alteration
a.UV light induced thymine- thymine
dimer
b.Alkylating agent cross linkage
3. Chain breaks
It is due to ionizing radiation,
radioactivity, oxidative free radicals.
4. Cross linkage

35. The mechanisms of DNA repair
1. Nucleotide excision repair (NER)
2. Mismatch repair (MMR)
3. Base excision repair (BER)
4. Homologous recombination repair
(HRR)
5. Non homologous end joining (NHEJ)

37. Double strand breaks

38. Transcription
 Synthesis of RNA from DNA
 Reading of DNA from 3’ to 5’
 Synthesis of RNA from 5’ to 3’
 RNA polymerase
 Promoter region regulates transcription
 Steps- Initiation , elongation, termination

39. Transcription

40. Transcription – RNA from DNA

41. Translation – RNA to proteins
Synthesis of proteins
mRNA – contains the message in the
form of codons
tRNA carries amino acids and
recognize codons with its anticodon
rRNA is involved in translation

43. Thank you