Questions and Answers
What does DNA polymerase need to start synthesis?
What is meant by 'semi-conservative' DNA replication?
Each newly replicated DNA double helix contains one intact parental strand and one newly synthesized daughter strand.
What is the origin of replication?
Unique DNA segments with short repeats recognized by origin-binding proteins.
What is a pre-replication complex (Pre-RC)?
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What are Okazaki fragments?
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How is the lagging strand formed?
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What is the role of DNA ligase?
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What are the two types of DNA polymerases in E.coli?
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RNA primers are required for DNA polymerase to initiate synthesis.
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What are the two steps in Okazaki fragment maturation?
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What is the 'end replication problem'?
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What is telomerase?
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What drugs target DNA replication machinery?
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Give examples of diseases associated with DNA replication defects.
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Study Notes
DNA Replication Concepts
- DNA replication is semi-conservative, meaning each new DNA helix contains one old and one new strand.
- Bidirectional replication initiates at replication origins and proceeds in both directions.
- Okazaki fragments are short DNA sequences synthesized discontinuously on the lagging strand.
- Replication origins are specific DNA segments recognized by origin-binding proteins, typically AT-rich.
- Replication forks are locations where DNA synthesis is actively occurring.
Requirements for DNA Synthesis
- DNA polymerase requires a template, nucleotides, an RNA primer for starting, and energy in the form of ATP.
RNA Primer and its Formation
- The RNA primer, roughly 10 nucleotides long, is synthesized by primase, an RNA polymerase that uses the parental DNA strand as a template.
Challenges Faced by DNA Polymerase
- DNA polymerase must address:
- Unwinding of the DNA helix.
- The need for a primer to initiate synthesis.
- Polarity issues during simultaneous synthesis of both strands, particularly solving lagging strand looping.
Pre-replication Complex (Pre-RC)
- The Pre-RC is a multi-protein complex formed around origins of replication, crucial for unwinding DNA by loading helicase (MCM) and requires kinase activation.
Unwinding and Stabilization Proteins
- DNA helicases are responsible for separating parental DNA strands and unwinding the double helix.
- Single-strand binding proteins (SSBPs) attach to DNA strands to maintain a single-stranded conformation and protect against nucleases.
DNA Polymerase Functionality
- DNA polymerase extends DNA by adding nucleotides to the growing strands from an RNA primer's 3' hydroxyl end.
Okazaki Fragment Maturation
- Maturation of Okazaki fragments involves:
- The removal of the RNA primer by DNA Pol I's exonuclease activity.
- Sealing gaps with DNA ligase which forms phosphodiester bonds.
Topoisomerase Role
- Topoisomerases relieve supercoiling tension that occurs ahead of the replication fork.
Telomerase Functionality
- Telomerase synthesizes telomeric repeat sequences at chromosome ends, maintaining chromosome integrity during replication; its activity is higher in stem and cancer cells.
Leading and Lagging Strand Synthesis
- The leading strand is synthesized continuously in the 3' to 5' direction toward the fork.
- The lagging strand is formed discontinuously away from the fork, resulting in the creation of Okazaki fragments.
Types of DNA Polymerases in E. coli
- Two types of polymerases:
- Distributive (DNA Pol I): Easily dissociates from the DNA strand.
- Processive (DNA Pol III): Remains attached for long stretches, enhancing replication efficiency.
Sliding Clamp Function
- The sliding clamp (PCNA in eukaryotes) increases the processivity of DNA polymerase by keeping it bound to the DNA template.
DNA Polymerases' Roles
- DNA Pol III is the primary enzyme for DNA replication in prokaryotes with high processivity.
- DNA Pol I is mainly involved in cleanup and replacing RNA primers with DNA.
Domains of DNA Polymerase
- DNA polymerase has two active sites:
- One for polymerization.
- A proofreading site (3' to 5' exonuclease activity) for correcting mistakes.
Discrimination Mechanism
- DNA polymerases discriminate base pairings using:
- Hydrogen bonding matching complementary bases.
- Geometric fit of A-T and G-C pairs in the active site.
End Replication Problem
- The end replication problem leads to telomere shortening with each cell division because the lagging strand cannot be fully synthesized.
- Telomerase reverses this by extending the lagging strand ends.
Drugs Targeting DNA Replication
- AZT: Inhibits HIV reverse transcriptase.
- Acyclovir: Targets viral DNA polymerase in herpes infections.
- Quinolones: Inhibit bacterial DNA gyrase.
Diseases Related to DNA Replication Defects
- Bloom Syndrome and Werner's Syndrome: Caused by mutations in helicases, leading to premature aging.
- Meier-Gorlin Syndrome: Linked to pre-replication complex mutations, causing dwarfism.
- FILS Syndrome: Resulting from DNA polymerase epsilon mutations, characterized by facial dysmorphism and immunodeficiency.
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Description
Explore key terms related to DNA replication such as semi-conservative, bidirectional, and Okazaki fragments. Understanding these concepts is crucial for grasping how genetic information is accurately copied during cell division. This quiz reinforces essential definitions and mechanisms involved in DNA replication.
