Eukaryotic DNA Replication and Repair Processes
Eukaryotic DNA Replication: Key Mechanisms
DNA molecules in eukaryotic cells are significantly larger than those in bacteria and are organized into complex structures. While essential features of replication are similar in eukaryotes and prokaryotes, eukaryotic initiation requires a protein complex. This interaction is regulated by various proteins involved in cell cycle control.
For instance, CDC6, similar to bacterial DnaA protein, functions to load helicases onto DNA near the origin of replication. However, the speed of movement of the replication fork in eukaryotes is only one-twentieth of that observed in E. coli.
Eukaryotic nuclear chromosomes involve various types of DNA polymerases in DNA replication:
- DNA Polymerase Alpha (Pol α): In association with DNA polymerase delta, Pol α lacks 3′-5′ exonuclease (proofreading) activity. It is primarily involved in synthesizing short RNA/DNA primers for Okazaki fragments.
- DNA Polymerase Delta (Pol δ): This polymerase elongates the primers. It is associated with a protein that stimulates it, known as Proliferating Cell Nuclear Antigen (PCNA). Pol δ possesses 3′-5′ exonuclease activity and performs the synthesis of both leading and lagging strands.
- DNA Polymerase Epsilon (Pol ε): Pol ε can replace Pol δ in some situations, such as DNA repair, and can also act at the replication fork.
Other proteins also play crucial roles in eukaryotic DNA replication:
- Replication Protein A (RPA): Functions equivalently to bacterial Single-Strand Binding (SSB) proteins.
- Replication Factor C (RFC): Acts as a clamp loader, facilitating the formation of replication complexes.
The termination of replication in linear eukaryotic chromosomes requires the synthesis of special structures called telomeres.
DNA Repair Mechanisms: Safeguarding Genetic Integrity
DNA can be damaged by various processes. The importance of DNA repair mechanisms lies in preventing a permanent change to the DNA nucleotide sequence, which is known as a mutation. The majority of mutations are deleterious, and only rarely are they advantageous.
Some types of DNA damage can be repaired by several different systems. Many repair systems are energy-intensive. DNA repair is largely possible because DNA has two complementary strands. An injury on one strand can be removed and accurately replaced using the undamaged complementary strand as a template.
Mismatch Repair (MMR)
For correcting wrong base pairing (mismatches), the repair system must somehow differentiate between the template strand and the newly synthesized strand. The cell achieves this by marking the DNA template with a methyl group.
In E. coli, this mechanism relies on DAM methylase, which methylates adenine residues at specific sites. The transient unmethylated state of the newly synthesized strand allows the cell to distinguish it from the template strand, enabling subsequent repair of mismatches. When a mismatch is located at the 5′ end of the unmethylated strand, that strand is unwound and degraded in the 3′-5′ direction, then replaced by new DNA.
Base Excision Repair (BER)
For Base Excision Repair (BER), specific DNA glycosylase enzymes exist in the cell that recognize DNA damage. Each glycosylase is typically specific for a particular lesion. While bacteria usually have only one type, eukaryotes possess at least four different DNA glycosylases.