DNA contains the instructions for building and operating every cell in an organism. Before a cell divides, this entire genetic material must be copied through DNA replication. Maintaining the integrity of this copy is paramount, as even small errors can have profound biological consequences. The central molecular machine responsible for this copying is the enzyme DNA Polymerase. This enzyme must operate with extreme accuracy, a fidelity it achieves through a built-in error-checking system.
DNA Replication and the Need for Accuracy
DNA replication requires the polymerase enzyme to read a template strand and synthesize a complementary new strand by adding nucleotides. Errors occur due to the immense scale of the task, as a human cell must copy approximately six billion base pairs during each division cycle. If the polymerase relied only on initial selectivity, it would incorporate a wrong base about once every 10,000 to 100,000 bases. This high initial error rate is biologically unacceptable and would lead to a flood of gene mutations. Cells have evolved multi-layered mechanisms to dramatically reduce this error frequency.
The Proofreading Function of DNA Polymerase
The primary mechanism for correcting initial mistakes is proofreading, performed by the DNA Polymerase itself. After adding a new nucleotide, the enzyme pauses momentarily to check the geometry of the newly formed base pair. Correct pairings, such as Adenine with Thymine, form a precise structure that fits perfectly into the enzyme’s active site. If an incorrect nucleotide is incorporated, the mismatched base pair distorts the local double helix structure. This distorted geometry causes the enzyme to stall its forward movement, signaling that the most recently added nucleotide is incorrect and must be removed.
Exonuclease Activity: The Correction Tool
Correction of the error detected during stalling is carried out by 3′ to 5′ exonuclease activity. DNA Polymerase possesses a secondary active site dedicated to this removal process. When the polymerase stalls due to a mismatch, the 3′ end of the newly synthesized strand shifts into the adjacent exonuclease domain. This domain removes the incorrectly incorporated nucleotide by cutting the phosphodiester bond. Once the faulty base is excised, the DNA strand shifts back to the polymerase active site to incorporate the proper nucleotide and resume synthesis in the 5′ to 3′ direction.
Secondary Defenses Against Gene Mutations
Proofreading is an immediate, but not final, check, as rare errors still escape this system. For these errors, the cell employs the Mismatch Repair (MMR) system, a complex of proteins that scans the newly synthesized DNA strand to recognize remaining mismatched bases or small insertion/deletion loops. A central challenge for MMR is distinguishing the correct template strand from the incorrect, newly synthesized strand containing the error. In bacterial systems, this distinction is made by detecting the temporary lack of methylation on the new strand, while human cells identify the new strand by searching for nicks. Once the faulty segment is identified, it is excised and replaced with the correct sequence by a repair DNA polymerase, reducing the final error rate of DNA replication to one in 10 billion bases.