DNA Replication

Fair warning, this lecture was terrible and I did not have the patience to make a good article. If anyone would like to contribute and improve the public knowledge base, please reach out or submit a pull request.

Cell Cycle Phases

M Phase: Cell splits
G1 Phase: Preparing for DNA copy
S Phase: DNA starts copying
G2 Phase: Preparing to split
G0 Phase: Resting, no replication

Mechanism

DNA synthesis is catalyzed by a nucleophilic substitution reaction. The 3'-OH of the growing polynucleotide chain acts as the nucleophile that attacks the incoming deoxynucleoside triphosphate (dNTP). After breaking the alpha-beta phosphoanhydride bond, the dNTP is incorporated, base-paired to a corresponding nucleotide.

The reaction is slightly exothermic, making it irreversible in vivo due to pyrophosphate hydrolysis.

DNA Polymerases

Alpha: Primase complex initiating Okazaki fragments
Delta/Epsilon: Analogous to E. coli's DNA polymerase III holoenzyme
Beta: Base excision repair
Gamma: Replicates mitochondrial DNA

Replisome Components

DNA helicase: Unwinds the DNA
Primase: Lays down RNA primers
DNA gyrase: Resolves supercoils

Initiation

ORC: A 6-protein complex binds at DNA starting points.
Activation: Proteins CDC6/CDC18 and CDT-1 bind to ORC.
Helicase Binding: These bindings allow another protein, MCM, to bind and unwind the DNA. Occurs in the G1 phase of the cell cycle.

Replication

Bidirectional, initiated at the origin, creating two replication forks. Newly synthesized DNA remains hydrogen-bonded to the template strand, following the semi-conservative replication model. This allows for error correction if a mistake occurs in copying the parental strand.

Leading Strand

DNA polymerase synthesizes new DNA in the 5' → 3' direction, maintaining continuous contact with the template, allowing for processivity.

Lagging Strand

Synthesis occurs in the 5' → 3' direction but in segments called Okazaki fragments. Primase lays down an RNA primer, followed by extension by DNA polymerase.

RNA primers are removed by DNA polymerase I's 5' → 3' exonuclease activity, and Okazaki fragments are glued back together by DNA ligase.

Fidelity Mechanisms

Base pairing geometry: Correct nucleotide insertion is facilitated by low Km, while incorrect nucleotides exhibit a high Km.
Proofreading: 3' → 5' exonuclease activity corrects erroneous insertions.

Tautomeric States

Cytosine can exist in high-energy tautomeric forms that can mispair but revert to their normal forms, creating an opportunity for error correction.