For BSc Msc

1. DNA REPLICATION
IN
PROKARYOTES

2. DNA REPLICATION
Contents
 Introduction
 DNA Structure
 Replication Model
 Enzymes involved in replication
 Mechanism of replication
 Replication in prokaryotes
 Conclusion
 Reference

3. What is replication?
DNA replication is the biological
process of producing two identical
copies of DNA from one
original DNA molecule.

4.  DNA replication occurs just before cell division(mitosis
and meiosis).
 It occurs during interphase.

5. Synthesis Phase (S Phase)
 S Phase During interphase of the cell cycle.
 DNA Replication take
place in the S Phase

6. DNA Structure
DNA has three main components
1. Deoxyribose (a pentose sugar)
2. Base (there are different ones)
3. phosphate

7. The Bases
There are divided into two groups
 Pyrimidines and purines
 Pyrimidines (made of one six member ring)
 Thymine
 adenine
 Purine (made of six member ring, fused to a 5 member ring)
 cytosine
 Guanine

9. DNA Double Helix and Hydrogen
Bonding
 Made of two strands of nucleotides that are joined
together by hydrogen bonding
 Hydrogen bonding occurs as a result of
complimentary base pairing
 Adenine and thymine pair up (2 bonds)
 Cytosine and guanine pair up(3 bonds)
 Each pair is connected through hydrogen bonding.

10. Modes of DNA Replication
Conservative
Semi-conservative
Dispersive

12.  Conservative
• Both parental strands stay together after DNA replication.
 Semiconservative
• The double-stranded DNA contains one parental and one daughter strand after
replication.
 Dispersive
• Parental and daughter DNA are interspersed in both strands after replication.

13. Enzymes of Replication
DNA Replication requires these enzymes:
1. DNA topoisomerase
2. Helicases
3. DNA Ligase
4. SSB proteins
5. RNA primase
6. DNA polymerase

14. 1. Helicases
Helicase use energy from the ATP to break the hydrogen bond holding the
base pair together.
This allows the two parental strands of DNA to begin unwinding and form
to replication forks.

15. 2. DNA topoisomerase
 Topoisomerases are enzymes that regulate the
overwinding or underwinding of DNA.
 It relieve stress ahead of replication fork.

16. 3. SSB Protein (single stranded DNA binding
protein)
When DNA was unwinded into two single strand by
helicase, it intend to form double stranded DNA and
keep the single state of DNA in replication.

17. 4. RNA Primase
Primase functions by synthesizing short
RNA sequences that are complementary
to a single-stranded piece of DNA, which
serves as its template.

18. DNA ligase
 DNA ligases close nicks in the phosphodiester
backbone of DNA. Biologically, DNA ligases are
essential for the joining of Okazaki fragments
during replication.

19. 5. DNA Polymerase
The enzyme responsible for DNA synthesis (replication)
and repair is called DNA Polymerase.

20. DNA polymerase
DNA polymerase I:
 It replace RNA primer with the DNA strand.
DNA Polymerase II:
 Polymerase II has 3’ to 5’ exonuclease activities and participate in DNA
repair.
DNA Polymerase III:
 It is responsible for leading and lagging strand synthesis.

21. Steps of DNA Replication
There are three steps involve in DNA replication.
1. Initiation
Replication begin at an the origin of the replication.
2. Elongation
New strands of DNA are synthesized by DNA Polymerase.
3. Termination
Replication is terminated differently in Prokaryotes and eukaryotes.

23. REPLICATION IN PROKARYOTES

24. DNA Replication in Prokaryotes
(E.coli) The genome of E.coli is replicated bi-directionally and
circular DNA. E.coli replicated is circular with no free
ends. Replication of DNA in E.coli is also known as
theta replication and it occurs in three steps.
1) Initiation
2) Elongation
3) Termination

25. INITIATION
 Helicase unwinding DNA forming a replication fork.
 To initiate replication process multiple replicative proteins must assemble
on these replication sites.

26. Elongation
 DNA polymerase III extends the RNA primer made
by primase.
 All DNA polymerases make DNA in 5’-3’ direction.
Two strand are formed.
I. Leading strand
II. Lagging strand

27. Leading strand synthesis
 On the template strand with 3’-5’ orientation, new
DNA is made continuously in 5’-3’ direction towards
the replication fork. The new strand that is
continuously synthesized in 5’-3’ direction is the
leading strand.

28. Leading & Lagging strand
DNA polymerase III synthesizes DNA for both leading
and lagging strands.
After DNA synthesis by DNA pol III, DNA polymerase I
uses its 5’-3’ exonuclease activity to remove the RNA
primer and fills the gaps with new DNA.
Finally DNA ligase joins the ends of the DNA fragments
together.

29. Lagging strand
 On the template strand with 5’-3’ orientation,
multiple primers are synthesized at specific sites by
primase (primosome complex) and DNA pol III
synthesizes short pieces of new DNA (about 1000
nucleotides long) new DNA is in 5’-3’ direction.
 These small DNA fragments that are discontinuously
synthesizes are called Okazaki fragments (named
after the discovered Reign Okazaki). The new strand
which is discontinuously synthesized in small
fragments is called the lagging strand.

31. Termination
 Once replication is complete, the two double
stranded circular DNA molecules remain interlinked.
Topoisomerase II makes double stranded cuts to
unlink these molecules.

32. Conclusion
 Helicase uncoils the DNA
 RNA primase adds short sequences of RNA to both strands (the
primer)
 The primer allows DNA polymerase III to bind and start replication
 DNA polymerase III adds nucleotides to each template strand in a
5'→3' direction
 These nucleotides are initially deoxyribonucleoside triphosphates but
they lose two phosphate groups during the replication process to
release energy
 One strand is replicated in a continuous manner in the same direction
as the replication fork (leading strand)
 The other strand is replicated in fragments (Okazaki fragments) in the
opposite direction (lagging strand)
 DNA polymerase I removes the RNA primers and replaces them with
DNA
 DNA ligase then joins the Okazaki fragments together to form a
continuous strand