DNA replication is the fundamental biological process by which a cell creates two identical copies of its deoxyribonucleic acid (DNA). This process is essential for cell division, allowing genetic information to be accurately passed from a parent cell to its daughter cells. It ensures the continuity of genetic material across generations. DNA replication is a highly coordinated and precise event, involving a complex machinery of specialized proteins and enzymes that duplicate the entire genome with remarkable accuracy.
Preparing the DNA for Copying
Before new DNA strands can be synthesized, DNA must be unwound and separated. The enzyme DNA helicase is responsible for unwinding the DNA double helix by breaking the hydrogen bonds that hold the two complementary strands together. This unwinding action creates a Y-shaped structure known as a replication fork, providing single-stranded DNA templates for the replication machinery.
As DNA helicase unwinds the helix, it introduces torsional stress, or supercoiling, in the DNA ahead of the replication fork. To prevent the DNA from tangling and potentially breaking, an enzyme called topoisomerase relieves this stress. Topoisomerase achieves this by making temporary nicks or breaks in the DNA strands, allowing the DNA to rotate and release the tension, and then resealing the breaks.
Once the DNA strands are separated, they are inherently unstable and have a tendency to re-anneal or form secondary structures. Single-strand binding proteins (SSBs) bind to these exposed single DNA strands. While not enzymes themselves, SSBs stabilize the separated strands, prevent them from coming back together, and protect them from degradation, ensuring the DNA remains accessible for the synthesis of new strands.
Initiating New Strand Synthesis
DNA polymerase cannot initiate DNA synthesis from scratch; it requires a pre-existing starting point. DNA primase is an enzyme that synthesizes short RNA sequences, called primers, which are complementary to the DNA template strand.
These RNA primers provide a free 3′-hydroxyl group. This specific chemical group is essential because DNA polymerase can only add new DNA nucleotides to an existing 3′-hydroxyl end. Therefore, primase’s role is to lay down these necessary RNA starting blocks, allowing DNA polymerase to begin its work of elongating the new DNA strand.
Building the DNA Strands
With the RNA primers in place, DNA polymerase synthesizes new DNA strands. This enzyme adds complementary DNA nucleotides to the 3′ end of the growing strand, using the original DNA strand as a template. DNA synthesis always proceeds in a 5′ to 3′ direction.
DNA polymerase is also equipped with a proofreading ability, which enhances the accuracy of DNA replication. As it synthesizes the new strand, the enzyme can detect and remove incorrectly paired nucleotides. If a wrong base is incorporated, the polymerase pauses, reverses direction, and uses an exonuclease activity to excise the mismatched nucleotide before resuming synthesis.
The antiparallel nature of the DNA double helix and the unidirectional activity of DNA polymerase lead to different modes of synthesis for the two new strands at the replication fork. One strand, known as the leading strand, is synthesized continuously in the same direction as the replication fork moves. The other strand, called the lagging strand, is synthesized discontinuously in short segments known as Okazaki fragments. This occurs because the lagging strand template runs in the opposite direction of the replication fork movement, requiring primase to repeatedly lay down new primers for each fragment.
Completing the Process
After the DNA polymerase has synthesized the new DNA strands, the RNA primers, which were initially laid down by primase, must be removed. These RNA segments are replaced with DNA nucleotides by a different DNA polymerase.
Once the RNA primers are replaced with DNA, small gaps or nicks remain in the sugar-phosphate backbone, particularly on the lagging strand between the newly synthesized Okazaki fragments. The enzyme DNA ligase plays a role in sealing these gaps. DNA ligase forms phosphodiester bonds, connecting the Okazaki fragments into a continuous, unbroken DNA strand, thereby completing the synthesis of the lagging strand.