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Questions and Answers
What is the primary purpose of DNA replication?
Which statement best describes the nature of DNA replication?
In which direction does DNA synthesis occur?
Which enzyme is primarily involved in unwinding double-stranded DNA prior to replication?
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What is required for DNA polymerase to initiate DNA synthesis?
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Where does DNA replication typically initiate?
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What components serve as precursors for DNA synthesis?
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What role does the primase enzyme play in DNA replication?
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What distinguishes the replication of the leading strand from the lagging strand?
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Which enzyme is responsible for synthesizing the final connections between DNA fragments?
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What initiates the formation of the replication fork during DNA replication?
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Which DNA polymerase is specifically involved in synthesizing the lagging strand?
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Which of the following describes single strand binding proteins (ssBP) in DNA replication?
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What is the directionality in which DNA polymerases synthesize a new strand?
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What is the function of exonucleases during DNA replication?
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What is one essential requirement for DNA polymerases to function in DNA replication?
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Which of the following components is NOT involved in the process of DNA replication?
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Which process ensures that each daughter DNA molecule contains one old strand and one new strand?
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What is the significance of the origin of replication (ori) in DNA replication?
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What type of molecular structure is primarily recognized by DNA helicases during the unwinding of DNA?
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In what manner does DNA synthesis occur on each of the two strands during replication?
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Which of the following statements about the direction of DNA polymerization is accurate?
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What are the characteristics of RNA primers used in DNA replication?
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How do DNA polymerases assist in the DNA replication process?
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What is the role of DNA ligase in DNA replication?
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What distinguishes the synthesis of the lagging strand during DNA replication?
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Which component is essential in maintaining the separation of DNA strands during replication?
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What is the primary function of exonucleases in DNA replication?
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Which of the following statements about DNA replication forks is true?
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What is the significance of the 3' to 5' reading direction of the template strand?
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Study Notes
DNA Replication
- DNA replication is the process of copying a DNA molecule into an identical copy.
- DNA replication occurs before cell division, so that the two daughter cells have identical DNA copies.
Semiconservative Replication
- Semiconservative replication refers to the mechanism by which each of the two resulting double-stranded DNA molecules contains one old strand and one newly synthesized strand from the original molecule.
- One old strand (parent strand) is conserved.
- One new strand is synthesized using free nucleotides found in the nucleus.
- Both strands serve as a template for DNA replication simultaneously.
Criteria for DNA Replication
- Replication is bidirectional.
- DNA synthesis occurs in the 5' to 3' direction.
- DNA replication requires a complex of enzymes and proteins.
Components of DNA Replication
- Template: Double-stranded DNA.
- Precursors: dATP, dGTP, dCTP, dTTP (deoxyribonucleotide triphosphates)
- Cofactors: Mg++, Mn++, ATP.
- Enzymes: DNA Polymerases, DNA helicases, DNA primase (a subunit of DNA polymerase α), Exonucleases.
- Proteins: Single-stranded DNA binding proteins (ssDB Proteins), Nick sealing enzyme DNA ligase.
DNA Polymerase
- DNA polymerase synthesizes a new DNA strand using the template strand.
- It reads the template strand in the 3' to 5' direction and synthesizes the new strand in the 5' to 3' direction.
- Deoxyribonucleotide triphosphates (dATP, dGTP, dCTP & dTMP) serve as precursors for the new strand.
- DNA polymerase cannot initiate DNA synthesis and it needs an RNA primer.
- There are five types of DNA polymerases in eukaryotes: α, ε, β, γ and δ.
Initiation of DNA Replication
- DNA replication begins at multiple DNA sites called origins of replication (ori).
- Origins of replication are rich in AT base pairs (consensus sequence).
- Multiple origins of replication begin simultaneously so replication occurs bidirectionally.
- DNA helicases break the hydrogen bonds between the two DNA strands, unwinding the double helix and creating a "replication bubble."
- Replication forks - The V-shaped structure at each end of the replication bubble.
- Single-stranded binding proteins (ssBP) hold the separated DNA strands apart to prevent them from rejoining.
Priming by Primase
- Primase is an RNA polymerase.
- Primase synthesizes a short RNA primer that provides a starting point for DNA polymerase.
- One RNA primer is required for the leading strand.
- Multiple primers are needed for the lagging strand.
Synthesis of Leading and Lagging Strands
- DNA polymerases are responsible for synthesizing both strands of DNA.
- The presence of many DNA polymerases (more than 20,000) reduces the time needed for replication.
Leading Strand
- Synthesized continuously.
- Replicated by DNA polymerase ε in the direction of the advancing replication fork.
Lagging Strand
- Synthesized discontinuously.
- Replicated by DNA polymerase δ in the opposite direction of the advancing replication fork.
- Synthesized in fragments called Okazaki fragments.
Excision of RNA Primers
- RNA primers are removed by exonucleases.
Ligation of DNA Fragments
- The final phosphodiester linkage between the 5'-phosphate group on one DNA chain and the 3'-hydroxyl group on the adjacent chain is catalyzed by DNA ligase.
- This reaction requires ATP.
Proofreading of Newly Synthesized DNA Strands
- DNA polymerases δ and ε have proofreading activity.
- They check for errors in base pairing and correct them, ensuring the accuracy of DNA replication.
DNA Replication
- DNA replication is the process of creating an identical copy of a DNA molecule.
- It occurs before cell division (mitosis) ensuring each daughter cell receives a complete set of DNA.
- DNA replication is semiconservative, meaning each new DNA molecule contains one original strand (parent strand) and one newly synthesized strand.
- Both DNA strands serve as templates for replication simultaneously.
- Replication proceeds bidirectionally, meaning it happens in both directions from the origin of replication.
- The direction of new DNA synthesis is always 5' to 3'.
Key Components of DNA Replication
-
DNA Polymerases: The main enzymes responsible for synthesizing new DNA strands.
- They require an existing RNA primer to begin replication.
- In eukaryotes, there are five DNA polymerases: α, ε, β, γ, and δ.
- DNA Helicase: Unwinds the double helix structure of DNA, separating the two strands.
- Single-stranded DNA binding proteins (ssBPs): Prevent the separated strands from re-annealing, keeping them accessible for replication.
- DNA Primase: Synthesizes short RNA primers that provide a starting point for DNA polymerase to attach to.
- Exonucleases: Remove RNA primers from the newly synthesized DNA strands.
- DNA Ligase: Seals the gaps between newly synthesized DNA fragments, creating a continuous strand.
Stages of DNA Replication
-
Initiation: Begins at specific sites called "origins of replication" (ori)
- Ori regions are rich in AT base pairs (adenine and thymine) due to their weaker bonding, making them easier to separate.
-
Unwinding and Separation:
- DNA helicase unwinds the double helix at the origin of replication, creating a "replication bubble".
- Two replication forks form at the ends of the bubble, and replication proceeds in opposite directions.
- Single-stranded binding proteins (ssBPs) prevent the separated strands from re-annealing.
-
Priming:
- Primase creates short RNA primers on both strands, providing starting points for DNA polymerase.
- The leading strand only needs one primer, while the lagging strand needs multiple primers.
-
Synthesis of Leading and Lagging Strands:
- Leading strand: Synthesized continuously in the 5' to 3' direction, following the replication fork movement.
- Lagging strand: Synthesized discontinuously in fragments (Okazaki fragments) in the opposite direction of fork movement.
-
Excision of RNA Primers:
- Exonucleases remove the RNA primers.
-
Ligation:
- DNA ligase joins the Okazaki fragments on the lagging strand and fills in any remaining gaps to make a continuous strand.
Proofreading Mechanism
- DNA polymerases (δ and ε) have a proofreading function.
- This mechanism helps ensure that the newly synthesized DNA strand is accurate and free of errors.
- DNA polymerase can backtrack (3' to 5' direction) and remove mismatched nucleotides.
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