Okazaki fragments are short sequences of DNA nucleotides approximately 150 to 200 base pairs long in eukaryotes which are synthesized discontinuously and after linked together by the enzyme DNA ligase to create the lagging strand during DNA replication. It is present on the template strand which read out new DNA synthesis away from the direction of replication fork movement. It is a building block for DNA synthesis of the lagging strand.
These were discovered in the 1960s by the Japanese molecular biologists Reiji and Tsuneko Okazaki, along with the help of some of their colleagues.
During DNA replication, the double helix is unwound and the complementary strands are separated by the enzyme DNA helicase, creating what is known as the DNA replication fork. Following this fork, DNA primase and then DNA polymerase begin to act in order to create a new complementary strand. Because these enzymes can only work in the 5’ to 3’ direction, the two unwound template strands are replicated in different ways.
One strand is the leading strand that undergoes a continuous replication process since its template strand has 3’ to 5’ directionality, allowing the polymerase assembling the leading strand to follow the replication fork without interruption. The other is the lagging strand that cannot be created in a continuous fashion because its template strand has 5’ to 3’ directionality, which means the polymerase must work backwards from the replication fork. This causes periodic breaks in the process of creating the lagging strand. The primase and polymerase move in the opposite direction of the fork, so the enzymes must repeatedly stop and start again while the DNA helicase breaks the strands apart.
Enzymes involved in the formation of Okazaki fragments:
- DNA polymerase
- DNA ligase I
- Flap endonuclease 1
- Dna2 endonuclease
Okazaki fragments in bacteria and in bacteriophage T4 are 1000–2000 nucleotides long, but are only about 100–300 nucleotides in eukaryotes.
Cellular DNA replication requires efficient copying of the double-stranded chromosomal DNA. The leading strand is elongated continuously in the direction of fork opening, whereas the lagging strand is made discontinuously in the opposite direction. The lagging strand needs to be processed to form a functional DNA segment. Genetic analyses and reconstitution experiments identified proteins and multiple pathways responsible for maturation of the lagging strand. In both prokaryotes and eukaryotes the lagging-strand fragments are initiated by RNA primers, which are removed by a joining mechanism involving strand displacement of the primer into a flap, flap removal, and then ligation. Although the prokaryotic fragments are ∼1200 nucleotides long, the eukaryotic fragments are much shorter, with lengths determined by nucleosome periodicity. The prokaryotic joining mechanism is simple and efficient. The eukaryotic maturation mechanism involves many enzymes, possibly three pathways, and regulation that can shift from high efficiency to high fidelity.