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Art connection

Illustration shows the replication fork. Helicase unwinds the helix, and single-strand binding proteins prevent the helix from re-forming. Topoisomerase prevents the DNA from getting too tightly coiled ahead of the replication fork. DNA primase forms an RNA primer, and DNA polymerase extends the DNA strand from the RNA primer. DNA synthesis occurs only in the 5' to 3' direction. On the leading strand, DNA synthesis occurs continuously. On the lagging strand, DNA synthesis restarts many times as the helix unwinds, resulting in many short fragments called “Okazaki fragments.” DNA ligase joins the Okazaki fragments together into a single DNA molecule.
A replication fork is formed when helicase separates the DNA strands at the origin of replication. The DNA tends to become more highly coiled ahead of the replication fork. Topoisomerase breaks and reforms DNA’s phosphate backbone ahead of the replication fork, thereby relieving the pressure that results from this supercoiling. Single-strand binding proteins bind to the single-stranded DNA to prevent the helix from re-forming. Primase synthesizes an RNA primer. DNA polymerase III uses this primer to synthesize the daughter DNA strand. On the leading strand, DNA is synthesized continuously, whereas on the lagging strand, DNA is synthesized in short stretches called Okazaki fragments. DNA polymerase I replaces the RNA primer with DNA. DNA ligase seals the gaps between the Okazaki fragments, joining the fragments into a single DNA molecule. (credit: modification of work by Mariana Ruiz Villareal)

You isolate a cell strain in which the joining together of Okazaki fragments is impaired and suspect that a mutation has occurred in an enzyme found at the replication fork. Which enzyme is most likely to be mutated?

The replication fork moves at the rate of 1000 nucleotides per second. DNA polymerase can only extend in the 5' to 3' direction, which poses a slight problem at the replication fork. As we know, the DNA double helix is anti-parallel; that is, one strand is in the 5' to 3' direction and the other is oriented in the 3' to 5' direction. One strand, which is complementary to the 3' to 5' parental DNA strand, is synthesized continuously towards the replication fork because the polymerase can add nucleotides in this direction. This continuously synthesized strand is known as the leading strand    . The other strand, complementary to the 5' to 3' parental DNA, is extended away from the replication fork, in small fragments known as Okazaki fragments , each requiring a primer to start the synthesis. Okazaki fragments are named after the Japanese scientist who first discovered them. The strand with the Okazaki fragments is known as the lagging strand    .

The leading strand can be extended by one primer alone, whereas the lagging strand needs a new primer for each of the short Okazaki fragments. The overall direction of the lagging strand will be 3' to 5', and that of the leading strand 5' to 3'. A protein called the sliding clamp    holds the DNA polymerase in place as it continues to add nucleotides. The sliding clamp is a ring-shaped protein that binds to the DNA and holds the polymerase in place. Topoisomerase prevents the over-winding of the DNA double helix ahead of the replication fork as the DNA is opening up; it does so by causing temporary nicks in the DNA helix and then resealing it. As synthesis proceeds, the RNA primers are replaced by DNA. The primers are removed by the exonuclease activity of DNA pol I, and the gaps are filled in by deoxyribonucleotides. The nicks that remain between the newly synthesized DNA (that replaced the RNA primer) and the previously synthesized DNA are sealed by the enzyme DNA ligase    that catalyzes the formation of phosphodiester linkage between the 3'-OH end of one nucleotide and the 5' phosphate end of the other fragment.

Once the chromosome has been completely replicated, the two DNA copies move into two different cells during cell division. The process of DNA replication can be summarized as follows:

  1. DNA unwinds at the origin of replication.
  2. Helicase opens up the DNA-forming replication forks; these are extended bidirectionally.
  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.
  5. Primase synthesizes RNA primers complementary to the DNA strand.
  6. DNA polymerase starts adding nucleotides to the 3'-OH end of the primer.
  7. Elongation of both the lagging and the leading strand continues.
  8. RNA primers are removed by exonuclease activity.
  9. Gaps are filled by DNA pol by adding dNTPs.
  10. The gap between the two DNA fragments is sealed by DNA ligase, which helps in the formation of phosphodiester bonds.


[link] summarizes the enzymes involved in prokaryotic DNA replication and the functions of each.

Prokaryotic DNA Replication: Enzymes and Their Function
Enzyme/protein Specific Function
DNA pol I Exonuclease activity removes RNA primer and replaces with newly synthesized DNA
DNA pol II Repair function
DNA pol III Main enzyme that adds nucleotides in the 5'-3' direction
Helicase Opens the DNA helix by breaking hydrogen bonds between the nitrogenous bases
Ligase Seals the gaps between the Okazaki fragments to create one continuous DNA strand
Primase Synthesizes RNA primers needed to start replication
Sliding Clamp Helps to hold the DNA polymerase in place when nucleotides are being added
Topoisomerase Helps relieve the stress on DNA when unwinding by causing breaks and then resealing the DNA
Single-strand binding proteins (SSB) Binds to single-stranded DNA to avoid DNA rewinding back.

Review the full process of DNA replication here .

Section summary

Replication in prokaryotes starts from a sequence found on the chromosome called the origin of replication—the point at which the DNA opens up. Helicase opens up the DNA double helix, resulting in the formation of the replication fork. Single-strand binding proteins bind to the single-stranded DNA near the replication fork to keep the fork open. Primase synthesizes an RNA primer to initiate synthesis by DNA polymerase, which can add nucleotides only in the 5' to 3' direction. One strand is synthesized continuously in the direction of the replication fork; this is called the leading strand. The other strand is synthesized in a direction away from the replication fork, in short stretches of DNA known as Okazaki fragments. This strand is known as the lagging strand. Once replication is completed, the RNA primers are replaced by DNA nucleotides and the DNA is sealed with DNA ligase, which creates phosphodiester bonds between the 3'-OH of one end and the 5' phosphate of the other strand.

Art connections

[link] You isolate a cell strain in which the joining together of Okazaki fragments is impaired and suspect that a mutation has occurred in an enzyme found at the replication fork. Which enzyme is most likely to be mutated?

[link] DNA ligase, as this enzyme joins together Okazaki fragments.

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Questions & Answers

what is cell?
V.S.Nikhil Reply
The smallest structure and functional unit
vinod
Hydra reproduce through which process
Saint Reply
which is smallest organ in our body
Techi
pineal gland
Himangshu
Yh in the ears...
Mozua
why you hand plam is sweating in everytime
Techi
who is the father of mycology
Sagar Reply
Heinrich Anton de Bary
Delissa
describe the similarities and differences between cytokinesis mechanism found in animal cells versus in plant cells
hiro Reply
what is life?
Techi
life is the existantce of indvidual human or animal.
R0se
thanks
Techi
are humans beings considered to have the eukaryotic cells
success Reply
yes.....
Delissa
eukaryotes are organisms that possess cells with a nucleus enclosed in a membrane, humans, and all complex organisms are eukaryotes.
Delissa
so humans and animals also have cell membranes.... cause I did this test prep and they said plants...I just want to be sure
success
and thank you for your reply it was helpful👍✌
success
eu= "perfect", "good", karyon= nut, amound, nucleus
Tiago
you're welcome. Plants are also eukaryotes.
Delissa
plants, like animals, possess a nucleus bound by a membrane.
Delissa
similarities and differences between cytokinesis mechanism found in animal cell vs cell division
Raymark Reply
what is the name of a male flower?
Ikeomu Reply
staminate means flower containing only stamen
Falak
what is the definition of evolution in a population?
Homero Reply
the slow changing of a species to adapt to any changes in the environment or how it feeds/hunts. im not good at explaining things lol.
Eclipse
the organ which is sensitive to light in euglena
Fatimah Reply
the organ which is sensitive to light in euglena is
Fatimah
all chlorophyll containing motile cells are sensitive to light
Himangshu
there is no more other chapter
Sandeep Reply
Give tow examples for nutritional deficiency Diseases-
Singampalli Reply
How does a plant cell look like
Sang Reply
in a sleepers form
David
what do you mean ? I could not understand
Gul
they have a regular shape and a large vacoule
Fatimah
I thought it looked like rectangle
Abrahán
a stage in mitosis wherein in spindle fibers begin to shorten to pu the sister chromatids away from each other towards the opposite ends of the cell
Earl Reply
a stage in interphase where chromosome s are duplicated
Earl
What is biodiversity
Sp Reply
Hmm
Hele
Name two secretions of Golgi apparatus
Daniel Reply

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Source:  OpenStax, Biology. OpenStax CNX. Feb 29, 2016 Download for free at http://cnx.org/content/col11448/1.10
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