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1.3. Dna Replication1

DNA replication takes place in the S phase of the cell cycle within the nucleus. It is a semiconservative process where each original DNA strand acts as a template to replicate a new complementary strand, resulting in two double-stranded DNA molecules each with one original and one new strand. Initiation involves enzymes unwinding and separating the DNA strands at the replication origin. RNA primers are laid down and DNA polymerase adds complementary nucleotides to extend the primers, replicating the DNA in the 5' to 3' direction on both the leading and lagging strands. DNA ligase seals together the Okazaki fragments on the lagging strand.

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38 views44 pages

1.3. Dna Replication1

DNA replication takes place in the S phase of the cell cycle within the nucleus. It is a semiconservative process where each original DNA strand acts as a template to replicate a new complementary strand, resulting in two double-stranded DNA molecules each with one original and one new strand. Initiation involves enzymes unwinding and separating the DNA strands at the replication origin. RNA primers are laid down and DNA polymerase adds complementary nucleotides to extend the primers, replicating the DNA in the 5' to 3' direction on both the leading and lagging strands. DNA ligase seals together the Okazaki fragments on the lagging strand.

Uploaded by

sabry tapia
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© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
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1.

3 Iniciación de la replicación
y control de la iniciación
DNA REPLICATION
DNA Replication
• New DNA strands are synthesized by using the existing (parent)
strands as templates in the formation of new, daughter strands that
are complementary to the parent strands.
• Could proceed by either a conservative or a semiconservative
mechanism:
– If a conservative mechanism were used, the two daughter
strands would form a new double-stranded DNA molecule and
the parent duplex would remain intact.
– If a semiconservative mechanism were used, the parent strands
would be permanently separated and each would form a duplex
molecule with the newly synthesized daughter strand base-
paired to it.
– Definitive evidence that duplex DNA is replicated by a
semiconservative mechanism came from a now classic
experiment conducted by M. Meselson and W. F. Stahl.
DNA is anti-parallel
• Two strands run
parallel to each other
but with opposite
alignments
(directions)
• McGraw-Hill DNA
• Why is being anti-
parallel an advantage
to the DNA
molecule?
DNA REPLICATION
• Semi-conservative =
each one of the
parent DNA strands
is passed to the
daugher DNA + one
new strand for each
• Semi-conservative
DNA (30 secs)
SNEAK PREVIEW:
DNA REPLICATION
PLAYERS (enzyme
review)
Question:
When and where does DNA Replication take
place?

Synthesis Phase (S phase)


• S phase in interphase of the cell cycle.
• Nucleus of eukaryotes
S
DNA replication takes phase
place in the S phase.
G1 interphase G2

Mitosis
-prophase
-metaphase
-anaphase
-telophase
DNA copying

• Each cell division cell must copy its entire DNA


• So each daughter cell gets a complete copy
• Rate of synthesis
– Bacteria = 1000 bases per second
– Mammals = 100 bases per second
• Problem - with a single replication origin in DNA
– Bacteria genome is 4 x 10E6. Takes 20 minutes to copy.
– Human is 3.2 x 10E9. Would take 10,000 times longer.
Initiation
• This process is initiated at particular points in the
DNA, known as "origins", which are targeted
by initiator proteins.
• In E. coli this protein is DnaA.
• In yeast, this is the origin recognition complex.
• Sequences used by initiator proteins tend to be "AT-
rich" (rich in adenine and thymine bases), because A-
T base pairs have two hydrogen bonds (rather than
the three formed in a C-G pair) and thus are easier to
strand-separate.
• Once the origin has been located, these initiators
recruit other proteins and form the pre-replication
complex, which unwinds the double-stranded DNA.
Fill out DNA Replication
Enzymes CHART
• Lots of enzymes are
needed to start each step
Enzyme Topoisomerase
also called DNA gyrase
• Unwinds double helix
Enzyme Enzyme

DNA
Topoisomerase
• Topoisomerase Youtube I and II (1:45)
• Topoisomerase Animation (2:16)
Enzyme Helicase:
separates (breaking H-
bonds) double helix at the
replication fork

YOU TUBE DNA replication (1:05)


Helicase
DNA Helicase
• The enzyme is unwinding the chain and
breaking the H-bonds between the
complementary base pairs (A-T, G-C).
Enzyme:
Primase
Helicase Primase
Helicase
= the enzyme
that makes RNA
nucleotides
into a primer
RNA Primer
• Nucleotides for the starting point
for DNA replication
• Short strands of RNA
DNA replication is initiated at many
points in eukaryotic chromosomes.
• Called Replication Bubbles
• They will eventually all meet to form whole
replicated strand
DNA Replication Bubble: DNA
duplicates in both directions
EM of DNA replication
Origins of Replication
• sites along the DNA molecule where
enzymes start the DNA replication - then
proceeds in both directions to form
“bubbles”
Replication Forks
Y-shaped regions of replicating DNA
molecules where new strands are
growing.
SSB’s
single strand binding proteins
• Stabilize the DNA strands as they
are being replicated
• Prevents rejoining of DNA strands
DNA Polymerases Require a
Primer to Initiate Replication

• DNA is synthesized from deoxyribonucleoside 5′-


triphosphate precursors (dNTPs).
• DNA synthesis always proceeds in the 5′→3′ direction
because chain growth results from formation of a
phosphoester bond between the 3′ oxygen of a growing
strand and the α phosphate of a dNTP.
• DNA polymerases cannot initiate chain synthesis de
novo; they require a short, preexisting RNA or DNA
strand, called a primer, to begin chain growth.
DNA Polymerases
• DNA Polymerase I
• Cuts off RNA primers and fills in with
DNA (between Okazaki fragments) –
lagging strand
• Can proofread
• DNA Polymerase III
• Elongates the strand by adding DNA
nucleotides on leading strand
• Also proofreads and corrects the
DNA strand
Anti-parallel strand builds in
the opposite direction (but
always in 5’ to 3’ direction)
Leading Strand Lagging Strand
• Template strand of • Other DNA strand
DNA • Forms short strands
• Continuous addition of Okazaki fragments
of nitrogenous bases (that will be joined
• in 5’ to 3’ direction later)
• McGraw-Hill Replication Fork • in the 5’ to 3’ direction
• DNA Replication You Tube (1:35)
• OKAZAKI FRAGMENTS
• The short strands of newly made DNA
fragments on the lagging strand are called
Okazaki fragments after the Japanese
Biochemist Reiji Okazaki.
•
Enzyme: DNA Ligase
a linking enzyme joins the strands

Example: joining two Okazaki


fragments together.

DNA ligase
Okazaki Fragment 1 Okazaki Fragment 2
5’ 3’

3’ Lagging Strand
5’
DNA LIGASE
is the enzyme that
joins the Okazaki
fragments (sugar -
phosphate backbone)
with covalent bonds

DNA REPLICATION
(look for ligase) 2:00
SUMMARY
DNA Replication (5:45) –
shows all the enzymes
• Summary Youtube of DNA replication (4:11)
• Good explanation of the 5’ to 3’ strands
and leading and lagging strands
Includes all your friendly
enzymes
• DNA Replication (3:56) Great animation
How Fast?

• Prokaryotic DNA polymerase can work


at about 1000 bases per second.

• Eukaryotic DNA polymerase can work at


about 50 bases per second.
Animation: DNA Replication
• DNA makes DNA
• *DNA with enzymes cartoons
All Together Now
• McGraw Hill Replication Fork animation
DNA Replication Easy Version (3:11)

• DNA Replication Youtube (7:48)

• Good CLICK and REVIEW


What if there is a mistake?

Typically
about one in
a billion
nucleotides
is incorrectly
paired
Proofreading

Initial base-pairing errors


are usually corrected by
DNA polymerase I.
Telomeres
• At the ends of each
chromosome is a
protective cap
called a telomere.
• Each time a cell
divides, the
telomeres are
snipped shorter,
Telomerase
• enzyme which adds DNA
sequence repeats
("TTAGGG" in all
vertebrates) to the 3' end of
DNA strands (an overhang)

• McGraw Hill Telomeres


Animation
Telomeres Added to ends of
chromosomes
Rutgers Telomere Animation
QUIZ
• Activity (choose your enzymes and
proteins)
MOVIE
• Media Showcase (cool animation)

• Replication Overview Movie

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