Whenever there is cell division, the genetic information, the cell comprises in the long strands of DNA, is copied by the enzymes referred to as the DNA polymerases. Each of the DNA strands renders a template which is used by the DNA polymerases to synthesise a complementary strand. One strand at the replication fork is continuously synthesised in the 5′ to 3′ direction (leading strand) while the (lagging strand) second strand is discontinuously synthesised in the 3′ to 5′ direction in short fragments which are referred to as the Okazaki fragments.
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Reiji Okazaki and Tuneko Okazaki, the Japanese molecular biologists, are said to be the ones who discovered these fragments in the 1960s, along with the contribution of some of their colleagues.
Okazaki Fragments Definition
Okazaki fragments are the short lengths of DNA that are produced by the discontinuous replication of the lagging strand.
The range of length of these fragments in the bacterial cells is about 1000-2000 nucleotides, while that in eukaryotic cells is approximately 100-200 nucleotides in length. The Okazaki fragments on the lagging strand are associated to generate a continuous new molecule of DNA.
Discovery of Okazaki Fragments
The Okazaki fragments were discovered by the pulse-labelling of the E.coli with 3H-thymidine in conditions that significantly reduced the rate of growth and division of cells. The E.coli were cultured at an optimum temperature of 37℃ for different generations in the existence of 14C-thymidine to label their DNA uniformly with 14C. Then the cells were cooled to 20℃ and pulse-labelled with 3H-thymidine for a duration of 10s for labelling of the nascent DNA in conditions wherein a decreased rate of DNA replication could disclose the existence of transient intermediates.
The time duration for the doubling of E.coli is about 40 minutes at a temperature of 37℃ and about 250 minutes at 20℃. In the pulse-chase experiments, a huge amount of unlabeled thymidine was supplemented to cells that were pulse-labelled for 10 s at a temperature of 20℃; incubation persisted for times suggested. The net cellular DNA then was isolated and fractionated by the sedimentation in alkaline sucrose gradients to denature it totally.
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The quantity of acid-insoluble radioactivity was quantified in each segment of the gradient, which could be soluble by treatment with deoxyribonuclease. In such conditions, the majority of the 3H-DNA first appeared as fragments about 50-5000 nucleotides approximately lengthwise, which then quickly became lengthened 3H-DNA fragments, ordered with a part as transient intermediates in the replication of DNA.
What Are Okazaki Fragments? – Overview
In eukaryotic cells, at the time of DNA replication, short single-stranded segments of DNA referred to as Okazaki fragments are synthesised, first on the lagging strand. These fragments originate from the 35-nucleotide-long-RNA-DNA primers. Once the Okazaki fragments are synthesised, the primers should be removed to permit the fragment to join into continuous lagging strands. A hindrance to elucidate Okazaki fragment processing is the dearth of methods that can examine the removal of primer directly in vivo.
At a replication fork of a replication bubble, the DNA synthesis is semi discontinuous. Both the daughter DNA molecules are synthesised at the replication fork. As the two DNA strands are antiparallel, one new strand should be synthesised in 5’ to 3’ direction, in the same direction as the fork approaches, but the other strand should be synthesised in a 3’ to 5’ direction overall, relative to the movement of the fork.
The enzymes which catalyze the addition of the deoxyribonucleotides to an increasing chain of DNA are referred to as DNA polymerases. If there is no 3’ to 5’ synthesising activity, how is the new strand aligned as 3’ to 5’ in the direction of the replication fork synthesised? The answer is the discontinuous synthesis of this strand, but the other strand is continuously synthesised.
Now, one of the template DNA strands is aligned as 3′ to 5′ at the replication fork, and as a result, it can be continuously copied by DNA polymerase, which extends the new DNA chain in a 5’ to 3’ direction. The new DNA chain is referred to as the leading strand with orientation 5′ to 3′ in the same direction as the movement of the fork. It extends from the origin of replication.
At the replication fork, the other template strand is oriented 5’ to 3’, as a result, copying it leads to the synthesis in a 3’ to 5’ direction with respect to the direction of the movement of the fork. This new chain of DNA is referred to as the lagging strand, which is discontinuously synthesised in a 3′ to 5′ direction. Such short fragments of DNA are joined together subsequently by the DNA ligase to produce an uninterrupted DNA strand. As the leading strand is continuously synthesised and the lagging strand is discontinuously synthesised, the complete process is called semi discontinuous.
What are Okazaki Fragments For?
Okazaki fragments are short sections of DNA formed at the time of discontinuous synthesis of the lagging strand during replication of DNA. It is essential as it allows for the synthesis of both the daughter strands required for cell division.
The role of Okazaki fragments is to permit the DNA polymerase to synthesise the lagging strands in the segments, as it is not correctly oriented for continuous synthesis.
Okazaki Fragments Formation
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As the DNA polymerase synthesises a part and then should wait for the helicase to open up more of the DNA helix upstream, the Okazaki fragments are formed on the lagging strand. Upon the opening up of the DNA by helicase, the primase gets in and puts down a new complementary RNA primer, which permits the DNA polymerase to associate the DNA and create the new Okazaki fragment.
Why are Okazaki Fragments Formed?
In most of the entities, the DNA acts as the genetic material. The DNA is double-stranded, comprising two DNA strands running antiparallel, which are linked by the hydrogen bonds. At the time of cell division, the entire DNA in the genome must be replicated, which doubles the DNA found in the original cell. In a semi-conservative mode, DNA replication takes place wherein one of the strands in the freshly created DNA (double-stranded) is the parent or the original strand. As a result, both the strands must act as templates in the replication of DNA. DNA polymerases are enzymes involved in the replication of DNA. They only synthesise DNA in the direction 5’ to 3’. But, due to the antiparallel nature of the double-stranded DNA, the synthesis of DNA must take place in either direction. Hence, the fragments take form at the time of synthesis of the lagging template strand.
Typically, the DNA polymerase adds the nucleotides in the direction 5’ to 3’. The enzymes are capable of continuously adding nucleotides to the growing strand on the leading strand. But, as the strand is running in the direction 5’ to 3’, the growth of the chain of the newly synthesised strand of DNA is put on hold when it arrives at the 5’ terminal of the strand. The synthesis of yet another DNA then starts at the replication fork.
This fork is the location on the DNA double-stranded wherein the unwinding starts, which is vital in synthesising the new strands of DNA on the parent strands. After the replication fork approaches the double-strand, the DNA polymerase can join nucleotides found on the lagging strand. But the synthesis gets halted when it arrives at the 5’ terminal of the RNA primer of the stretch of DNA that is already synthesised. Consequently, the synthesis of DNA at the lagging strand is not continuous, while the resultant stretches of DNA are the Okazaki fragments.
Okazaki Fragments Function
Okazaki fragments are primarily involved in enabling the DNA polymerase in synthesising the lagging strand, even though it is oriented in the opposite direction. A type of DNA polymerase, DNA polymerase I, arrives and removes the RNA primers, replacing them with DNA. The Okazaki fragments should be attached into one continuous strand once replication occurs. This is achieved by the DNA ligase that seals the sugar-phosphate backbone of the Okazaki fragments. This enables the replication of two continuous, identical daughter DNA strands.
Before cell division occurs, it is vital to replicate DNA wherein one parent cell splits to produce two daughter cells, which makes sure that both the daughter cells obtain the same genetic material. Cell division in unicellular entities could be a mode of asexual reproduction, while cell division in multicellular entities is vital for the repair and growth of the entity and to give rise to cells required for sexual reproduction.
This was a brief on Okazaki fragments. For more related concepts, visit BYJU’S NEET.
Also see:
- Double Helix Structure of DNA
- When does DNA Copying occur?
- Importance of Cell Division
- RNA splicing
- What are Telomeres?
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