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DNA

Replication
By: Micah Ezra N. Chang
The Big Questions
1. Why do we need to study DNA Replication?

2. What is DNA Replication?

3. How can we apply our knowledge about DNA


Replication to our classroom or daily life?
Why do we need to
study DNA Replication?
The Why do we need to study DNA Replication?

1. To understand cancer
2. To understand aging

3. To understand diseases related to DNA repair


● a) Bloom’s Syndrome
● b) Xeroderma Pigmentosum
● c) Werner’s Syndrome
What is DNA
Replication?
TOPICS

Basics of DNA
01 Replication
DNA Replication
03 in Eukaryotes
DNA Replication
02 in Prokaryotes
01: Basics of DNA
Replication
Learning Objectives:
• Explain how the structure of DNA reveals the
replication process
• Describe the Meselson and Stahl experiments
01: Basics of DNA Replication

DNA structure
● A DNA (Deoxyribonucleic acid) molecule looks like
a twisted ladder. Its shape is called a double helix.
A helix is a shape that twists.
● The two sides of the DNA ladder are made of sugar
molecules alternating with phosphate molecules.
● The rungs of the DNA molecule are made of
chemical building blocks called bases. The four
bases found in DNA are adenine (A), thymine (T),
cytosine (C), and guanine (G).
01: Basics of DNA Replication

● Before mitosis the amount of DNA doubles.


● DNA replication is the process of a DNA molecule
making a copy of itself.
● DNA replication occurs before mitosis begins and
before the first division of meiosis.
DNA replication ensures that each
daughter cell has an exact copy of the
DNA from the parent cell.
01: Basics of DNA Replication

● DNA replication results in one DNA molecule


becoming two daughter molecules—each an exact
copy of the original molecule.

● DNA replication requires:


○ A set of proteins and enzymes
■ DNA polymerase, also known as DNA pol
● DNA pol adds nucleotides one-by-
one to the growing DNA chain that is
complementary to the template
strand.
○ Energy in form of ATP
01: Basics of DNA Replication

● The double-helix model suggests that the two


strands of the double helix separate during
replication, and each strand serves as a template
from which the new complementary strand is
copied.
● What was not clear was how the replication took
place.
● There were three models suggested
○ Conservative
○ semi-conservative
○ dispersive.
01: Basics of DNA Replication

● Conservative replication:
– The parental double helix remains intact;
– both strands of the daughter double helix are newly synthesized

It would leave the original template DNA strands intact and would
produce a copy composed of entirely new DNA base pairs.
01: Basics of DNA Replication

● Semiconservative replication:
–It would produce two copies that each contained one of the original
strands, and one entirely new copy.
01: Basics of DNA Replication

● Dispersive replication:
–At completion, both strands of both double helices contain both original
and newly synthesized material.

Dispersive replication would produce two copies of the DNA, both


containing a mixture of old and new DNA base pairs.
01: Basics of DNA Replication

The deciphering of the structure of DNA by Watson and Crick in 1953


suggested that the semiconservative model was correct (as Watson and Crick
pointed out in a sly one-line concluding sentence to their seminal paper).
This was soon verified by Meselson-Stahl experiment.
01: Basics of DNA Replication

Meselson-Stahl experiments
○ Experiment allowed
differentiation of parental and
newly formed DNA.
01: Basics of DNA Replication

● The steps of DNA replication can be summarized


into four:

Step 1: Replication Fork Formation


Step 2: Primer Binding
Step 3: Elongation
Step 4: Termination
02: DNA
Replication in
Prokaryotes
Learning Objectives:
• Explain the process of DNA replication in
prokaryotes
• Discuss the role of different enzymes and
proteins in supporting this process
02: DNA Replication in Prokaryotes

● DNA replication in Prokaryotes requires:


○ Three main types of polymerases are known:
DNA pol I, DNA pol II, and DNA pol III.
■ DNA pol I is an important accessory
enzyme in DNA replication,
■ along with DNA pol II, is primarily
required for repair.
■ DNA pol III is the enzyme required for
DNA synthesis.
02: DNA Replication in Prokaryotes

Step 1: Replication Fork


Formation
1. DNA unwinds at the origin of
replication.
How does the replication machinery
know where to begin? It turns out that
there are specific nucleotide sequences
called origins of replication where
replication begins.

2. Helicase opens up the DNA-


forming replication forks;
these are extended
bidirectionally.
02: DNA Replication in Prokaryotes

Step 1: Replication Fork


Formation
3. Single-strand binding
proteins coat the DNA around
the replication fork to prevent
rewinding of the DNA.
4. Topoisomerase binds at
the region ahead of the
replication fork to prevent
supercoiling
02: DNA Replication in Prokaryotes

Step 2: Primer Binding

5. Primase synthesizes RNA


primers complementary to the
DNA strand.
6. DNA polymerase III starts
adding nucleotides to the 3'-OH
end of the primer.
02: DNA Replication in Prokaryotes

Step 2: Primer Binding


02: DNA Replication in Prokaryotes

Step 3: Elongation

7. Elongation of both the


lagging and the leading strand
continues.
02: DNA Replication in Prokaryotes

Step 4: Termination

8. RNA primers are removed


by exonuclease activity.
9. Gaps are filled by DNA pol I
by adding dNTPs.
10. The gap between the two
DNA fragments is sealed by
DNA ligase, which helps in the
formation of phosphodiester
bonds.
02: DNA Replication in Prokaryotes
03: DNA
Replication in
Eukaryotes
Learning Objectives:
• Discuss the similarities and differences
between DNA replication in eukaryotes and
prokaryotes
• State the role of telomerase in DNA replication
03: DNA Replication in Eukaryotes

The DNA Replication in Eukaryotes:


Step 1: Replication Fork Formation
Step 2: Primer Binding
Step 3: Elongation
Step 4: Termination
03: DNA Replication in Eukaryotes
Eukaryotic DNA is bound to basic proteins known
as histones to form structures called
nucleosomes. Histones must be removed and
then replaced during the replication process,
which helps to account for the lower replication
rate in eukaryotes.

The number of DNA polymerases in eukaryotes


is much more than in prokaryotes: 14 are known,
of which five are known to have major roles
during replication and have been well studied.
They are known as pol α, pol β, pol γ, pol δ, and
pol ε.
02: DNA Replication in Prokaryotes

At the origin of replication, a


pre-replication complex is made
with other initiator proteins
Step 1: Replication Fork
Formation
1. DNA unwinds at the origin of
replication.
2. Helicase opens up the DNA-
forming replication forks;
these are extended
bidirectionally.
02: DNA Replication in Prokaryotes

Step 1: Replication Fork


Formation
3. Single-strand binding
proteins coat the DNA around
the replication fork to prevent
rewinding of the DNA.
4. Topoisomerase binds at
the region ahead of the
replication fork to prevent
supercoiling
02: DNA Replication in Prokaryotes

Step 2: Primer Binding

Primers are formed by the enzyme


primase, and using the primer, DNA pol
can start synthesis.
Three major DNA polymerases are then
involved: α, δ and ε.
● DNA pol α adds a short (20 to 30
nucleotides) DNA fragment to the
RNA primer on both strands, and
then hands off to a second
polymerase.
02: DNA Replication in Prokaryotes

Step 3: Elongation

● While the leading strand is


continuously synthesized by the
enzyme pol ε, the lagging strand is
synthesized by pol δ
● A sliding clamp protein known as
PCNA (proliferating cell nuclear
antigen) holds the DNA pol in place
so that it does not slide off the DNA.
02: DNA Replication in Prokaryotes

Step 4: Termination

• The primer RNA is then removed by


RNase H (AKA flap endonuclease)
and replaced with DNA nucleotides.
• The Okazaki fragments in the lagging
strand are joined after the
replacement of the RNA primers with
DNA.
• The gaps that remain are sealed by
DNA ligase, which forms the
phosphodiester bond.
02: DNA Replication in Prokaryotes

Telomere replication
Unlike prokaryotic chromosomes,
eukaryotic chromosomes are linear.
The DNA at the ends of the
chromosome thus remains unpaired,
and over time these ends, called
telomeres, may get progressively
shorter as cells continue to divide.
02: DNA Replication in Prokaryotes

Telomere replication
● Telomeres comprise repetitive
sequences that code for no
particular gene.
● Telomeres protect the genes from
getting deleted as cells continue to
divide. The telomeres are added to
the ends of chromosomes by a
separate enzyme, telomerase.
● The telomerase enzyme contains a
catalytic part and a built-in RNA
template
02: DNA Replication in Prokaryotes

Telomere replication
02: DNA Replication in Prokaryotes

Telomere replication
02: DNA Replication in Prokaryotes

Telomere replication
02: DNA Replication in Prokaryotes

Telomere replication
02: DNA Replication in Prokaryotes

Telomerase and Aging


● Cells that undergo cell division continue to have their telomeres shortened
because most somatic cells do not make telomerase. This essentially means
that telomere shortening is associated with aging.
● In 2010, scientists found that telomerase can reverse some age-related
conditions in mice. Telomerase reactivation in these mice caused extension
of telomeres, reduced DNA damage, reversed neurodegeneration, and
improved the function of the testes, spleen, and intestines. Thus, telomere
reactivation may have potential for treating age-related diseases in
humans.
02: DNA Replication in Prokaryotes

Cancer is characterized by uncontrolled cell division of abnormal cells. The


cells accumulate mutations, proliferate uncontrollably, and can migrate to
different parts of the body through a process called metastasis. Scientists have
observed that cancerous cells have considerably shortened telomeres and that
telomerase is active in these cells. Interestingly, only after the telomeres were
shortened in the cancer cells did the telomerase become active. If the action of
telomerase in these cells can be inhibited by drugs during cancer therapy, then
the cancerous cells could potentially be stopped from further division.
How can we apply our
knowledge about DNA
Replication to our
classroom or daily life?
How can we apply our knowledge about DNA
Replication to our classroom or daily life?

● Knowing our Telomeres Substantially Improves


Quality of Life
● 5 ways to encourage telomere lengthening and delay
shortening
● 1. Maintain a healthy weight.
● 2. Exercise regularly.
● 3. Manage chronic stress.
● 4. Eat a telomere-protective diet.
● 5. Incorporate supplements.
Thanks!
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