WHAT IS DNA REPLICATION?
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No molecular structure has gained more worldwide recognition than the DNA double helix.
James Watson and Francis Crick wrote the famous Nature paper in 1953 entitled, ​Molecular
Structure of Nucleic Acids​, which ends with, “It has not escaped our notice that the specific
pairing we have postulated immediately suggests a possible copying mechanism for the genetic
material.” Since then, many scientists have focused on researching the mechanism of DNA
replication.
Figure 1
ACTIVITY ONE
Using Figure 1 above as a reference. Double
click the image to the right and place the
circle around the 3’ (three prime) carbon and
the 5’ (five prime) carbon in the DNA
nucleotide shown in the diagram on the right.
Primers are used in numbering carbons
on the sugar (deoxyribose) portion of the
nucleotide to distinguish them from the
nitrogenous base carbons.
DNA PRIMERS
Double-stranded DNA is composed of two antiparallel strands! Each DNA strand has
directionality. The two sugar-phosphate backbones run in opposite 5’ 3’ directions from each
other.
1. Use the figures above to answer the following question. What group is attached to
the 3’ carbon? T
​ he Hydroxyl Group
2. Use the figures above to answer the following question. What group is attached to
the 5’ carbon? T
​ he Phosphate Group
Figure 2
3. How many nucleotides are present in Figure 2? S
​ ix.
4. How many strands of DNA are shown in Figure 2? ​Two.
5. What is the difference between the strand on the left and the strand on the right?
They are going in opposite directions. The strand on the left is going from 5’ to 3’
and the strand on the right is going from 3’ to 5’.
6. What types of bonds connect the nucleotide 1,2, and 3 together? ​Covalent bonds
connect the nucleotides in a single strand of DNA.
7. What types of bonds connect nucleotide 1 to nucleotide 6? H
​ ydrogen bonds
connect nitrogenous base pairs of a double strand of DNA.
THE REPLICATION FORK
DNA replication begins at specific sites called origins of replication. A eukaryotic chromosome
may have hundreds or even a few thousand replication origins. Proteins that start DNA
replication attach to the DNA and separate the two strands, creating a replication bubble. These
proteins are called ​primers​​ and you will learn more about them below. At each end of the
replication bubble is a Y-shaped region where the parental strands of DNA are being unwound.
This region is referred to as the replication fork.
Figure 3
3. Reviewing Figure 3 and reflection on the the double-helix structure of DNA, what would
be the first step of DNA replication? U
​ nwinding or separating the DNA.
4. Why do you think multiple replication bubbles form during the process of DNA
replication? ​The replication process would be too slow if replication occurs at a single
bubble.
Figure 4: ​3-D animation shows how DNA is replicated at the molecular level
HELICASE
During DNA replication, both strands of the double helix act as templates for the formation of
new DNA molecules. Copying occurs at a localized region called the replication fork, which is a
Y shaped structure where a multi-enzyme complex synthesizes new DNA strands. One of the
enzymes found in that enzyme complex is called​ helicase. ​Helicase is the protein structure that
is ​blue ​in Figure 4.
5. Studying the image above, what is the function of the enzyme helicase? ​The function
of helicase is to unwind the double stranded DNA helix.
6. What type of bond is being broken by helicase? H
​ elicase is breaking the hydrogen
bonds between the nitrogenous bases of the 2 DNA strands
DNA POLYMERASE: Continuous replication
The enzyme helicase spins the incoming DNA to unravel it: at ten thousand RPM in the case of
bacterial systems. The separated strands are called three prime and five prime, distinguished by
the direction in which their component nucleotides join up. The 3' DNA strand, also known as
the ​leading strand​, is diverted to a ​DNA polymerase. ​The DNA polymerase enzyme catalyzes
the synthesis of new DNA by adding nucleotides to a preexisting chain. Note, new DNA can
elongate only in the 5’ 3’ direction. DNA polymerase uses the leading strand as a continuous
template for the synthesis of the first daughter DNA helix.
7. In your own words, describe the function of DNA polymerase. ​DNA polymerase adds in
the new template of nucleotides to the original strand of DNA separated by helicase.
8. Nucleotides are added at an approximate rate of 50 nucleotides per second in
eukaryotic cells. The human genome contains 6.4 billion nucleotides (3.2 billion base
pairs), which must be copied. Calculate the length of time in days that it would take to
copy the human genome. Show all calculations including units. A
​ pproximately 1,500
days.
9. As a new nucleotide is added to the growing DNA strand, which part of the new
nucleotide forms a bond with the 3’ OH group? (Hint study figure 1 and figure 2 for help)
The phosphate group.
ACTIVITY TWO
Double click the image and use one arrow to
indicate the directionality of the newly
replicated​ leading strand​​ of DNA.
Note: The 3’ OH group is essential for adding
a new nucleotide to the growing DNA strand.
If this group is not present – for example, if
there is a 3’ H instead of a 3’ OH – then DNA
synthesis cannot continue. This is the basis
for the development of the Sanger
sequencing method, which is used in
determining the sequence of nucleotides.
Teacher’s Note: Remember, DNA is antiparallel so adding in a 5’-3’ will be going the opposite
direction of the original strand. Reference the image below for a better understanding.
DNA POLYMERASE: Discontinuous replication
Replication occurs on both sides of the replication fork simultaneously. The other half of the
DNA double helix, known as the ​lagging strand,​ has the opposite 3' to 5' orientation and
consequently requires a more complicated copying mechanism. As it emerges from the
helicase, the lagging strand is organized into sections called ​Okazaki fragments.​ These
Okazaki fragments and are usually 100-200 nucleotides long in eukaryotic cells. These are then
presented to a second DNA polymerase enzyme in the preferred 5' to 3' orientation. These
sections are then effectively synthesized backward. The DNA polymerase must move away
from the fork instead of toward the fork, as it did in the leading strand. This is to accommodate
the 5’ 3’ synthesis of DNA, short fragments are made on the second strand, which is referred to
as the lagging strand.
ACTIVITY THREE
Double click the image and use one arrow to
indicate the directionality of the newly
replicated ​lagging strand​​ of DNA.
TEACHERS NOTE: This is the direction
PRIMASE & LIGASE
Remember there is an enzyme complex involved in DNA replication. Specifically, there are
multiple enzymes involved in copying the lagging strand. ​Primase​ adds R
​ NA primers​ onto the
leading and lagging strands of DNA. In the leading strand, it acts as a starting point to alert DNA
polymerase were to add a short row of DNA bases. In the lagging strand, this allows synthesis
of Okazaki fragments from 5' to 3'. How? The primase acts as a temporary stop signal, briefly
halting the progression of the replication fork during DNA replication.​ In the end, ​DNA ligase​​ is
an enzyme that repairs irregularities or breaks in the backbone of double-stranded DNA
molecules. It has three general functions: It seals repairs in the DNA, it seals recombination
fragments, and it connects Okazaki fragments. The actual process of joining the Okazaki
fragments together is more complex and involves several other proteins. DNA replication is
considered to be a semi-discontinuous process.
10. What is the function of primase enzyme in the lagging versus the leading strand?
Primase act as the start point of DNA replication. It tells the DNA polymerase where to
start adding in the new strand of complementary base pairs. It does this by adding in an
RNA primer.
11. What is the function of the ligase enzyme in DNA replication? ​Ligase acts like the
glue for DNA replication, sealing in fragments to have a continuous new strand of DNA.
12. Why is DNA replication considered to be a semi-discontinuous process? B
​ ecause the
process has new one strand that is added continuously, the leading strand, and one new
strand that is added discontinuously, the lagging strand.
13. DNA is often described as being semiconservative. In your own words describe what
that means. D
​ NA replication is a semi-conservative process, because when a new
double-stranded DNA molecule is formed by one strand will be from the original template
molecule and one strand will be newly synthesised
14. How do the two new strands of DNA compared to the original (parental) strand?​ It is
moving in the opposite direction since DNA is antiparallel and it contains the
complementary base pairs of the original parent DNA strand.
15. What is the relationship of DNA replication to cell division? ​DNA replication is the
process by which cells make make a copy of DNA for the 2 new daughter cells during S
phase of Interphase.
THE BIG IDEA
Each time one of your cells divide, it must duplicate its DNA genome so that the new cell will
have a replica of DNA contained in the original cell. This process of duplicating the
double-stranded DNA is called replication. Because the nucleotide sequence of one strand of
DNA is complementary to the sequence of the opposite strand – during DNA replication, the two
strands of the original DNA are separated, and two different DNA polymerase enzymes use
every single strand as a template for the synthesis of the second, complementary strand. This
process is known as semiconservative DNA replication – meaning that each new cell receives a
double-stranded DNA composed of one old strand of DNA and one newly-synthesized strand.
For a better realistic video of the process of DNA replication, watch this video.​DNA Replication
Video
NEXT STEPS: WHAT TO ADD IN YOUR NOTEBOOK
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Still having trouble? Watch this simplified video explaining DNA replication by
CLICKING HERE​.
For an even more simplified video, watch this Amoeba Sister Video on DNA replication
by ​CLICKING HERE​.
Create a graphic organizer that list all 4 enzymes involved in DNA replication and their
function.
Summarize the steps of DNA replication starting with Helicase and ending with Ligase.
Do not forget to use your vocabulary words. (quick tip: underline major vocabulary
words in your summary)