DNA holds the genetic instructions that guide the development, functioning, growth, and reproduction of all known organisms. For life to persist and for cells to divide, this genetic instruction set must be accurately duplicated. This process is known as DNA replication. It ensures that each new cell receives a complete and identical copy of the genetic material, making it central for biological inheritance and the continuity of life.
Preparing the DNA for Copying
DNA replication begins with the unwinding of the double helix structure. An enzyme called helicase “unzips” the DNA molecule by breaking the hydrogen bonds between the complementary base pairs, creating a Y-shaped replication fork. This unwinding process can introduce tension in the DNA ahead of the replication fork. Topoisomerase enzymes work to relieve this supercoiling, preventing the DNA from becoming tangled and damaged.
As the DNA strands separate, they are kept from re-annealing by single-strand binding proteins (SSBPs). These proteins coat the single strands, stabilizing them and keeping them accessible for the replication machinery. Since DNA polymerase cannot start a new strand from scratch, a short RNA segment called a primer is synthesized by an enzyme called primase. This RNA primer provides a starting point for DNA synthesis.
Building the New DNA Strands
Once the DNA template strands are prepared and primers are in place, DNA polymerase enzymes begin to add new DNA nucleotides. DNA polymerase can only add nucleotides in one direction, from the 5′ end to the 3′ end of the new strand. This directional limitation means that one of the new DNA strands, called the leading strand, can be synthesized continuously as the replication fork opens. A single RNA primer is sufficient to initiate synthesis on the leading strand.
In contrast, the other strand, known as the lagging strand, is synthesized discontinuously. Because its template runs in the opposite direction, DNA polymerase must synthesize this strand in short segments, moving away from the replication fork. These short fragments are called Okazaki fragments. Each Okazaki fragment requires its own RNA primer to start synthesis.
Finishing and Sealing the New DNA
After the DNA polymerase has synthesized the new DNA segments, particularly the Okazaki fragments on the lagging strand, the RNA primers that initiated these segments must be removed. Different types of enzymes, such as nucleases, then remove these RNA primers and fill the resulting gaps with DNA nucleotides.
Following primer removal, the gaps left behind are filled in with DNA by another DNA polymerase. The final step in creating a continuous DNA strand involves DNA ligase. This enzyme forms phosphodiester bonds, effectively sealing the nicks or breaks between the newly synthesized DNA fragments, especially joining the Okazaki fragments on the lagging strand into one complete, unbroken strand.
Maintaining DNA Accuracy
The process of DNA replication includes built-in mechanisms to ensure high accuracy. DNA polymerase itself has a “proofreading” ability, which allows it to check each newly added nucleotide. If an incorrect nucleotide has been incorporated, the polymerase can remove it and replace it with the correct one before continuing synthesis. This proofreading function significantly reduces the number of errors during replication.
Even with proofreading, a small number of errors can still occur. For these, cells employ additional repair mechanisms, such as mismatch repair. Mismatch repair systems identify and correct incorrectly paired bases that were missed by proofreading, further safeguarding the integrity of the newly synthesized DNA. These mechanisms are important for maintaining genetic stability and preventing mutations that could lead to various health issues.