Replication

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Questions and Answers

What is the result of DNA replication in semiconservative replication?

One new DNA molecule with one original strand and one newly synthesized strand (correct)

Two completely new DNA molecules with no original strands

Four new DNA molecules with two original strands and two newly synthesized strands

One original DNA molecule with no changes

What is the role of 15NH4Cl in the experiment by Meselson and Stahl?

To separate the two original DNA strands

To slow down DNA replication

To label newly formed nitrogen compounds, including purine and pyrimidine nucleobases, with a heavy isotope of nitrogen (correct)

To synthesize new polynucleotide chains

What is the technique used to analyze the extracted DNA in the experiment by Meselson and Stahl?

Density-gradient centrifugation (correct)

Electrophoresis

Spectroscopy

Chromatography

What is the direction of DNA synthesis in most organisms?

Bidirectional Signup and view all the answers

What is the structure formed by the advance of the two replication forks in opposite directions?

All of the above Signup and view all the answers

How many origins of replication exist in eukaryotes?

Several Signup and view all the answers

What is the direction of synthesis of new polynucleotide chains?

5' to 3' Signup and view all the answers

Why do new polynucleotide chains grow in the 5' to 3' direction?

Due to the nature of the reaction of DNA synthesis Signup and view all the answers

What is the significance of the 3'-hydroxyl on the sugar of the last nucleotide added to a growing chain?

It determines the direction of DNA synthesis Signup and view all the answers

What is the term used to describe the growth of both new polynucleotide chains in bidirectional DNA replication?

Net chain growth Signup and view all the answers

What are the two forms of naturally occurring DNA?

Single-stranded and double-stranded Signup and view all the answers

What is the primary challenge in separating the two DNA strands?

Unwinding the double helix Signup and view all the answers

Why must the cell protect the unwound portions of DNA?

To prevent nucleases from attacking single-stranded DNA Signup and view all the answers

In which direction are the two antiparallel strands synthesized?

From the 5' to the 3' end Signup and view all the answers

What is the purpose of the cell's quality control mechanism during replication?

To ensure the correct base is added to the growing polynucleotide chain Signup and view all the answers

Why is the replication of double-stranded DNA complex?

Because it allows for a high degree of fine-tuning and fidelity in replication Signup and view all the answers

Which organism was primarily used to study the details of DNA replication?

Escherichia coli Signup and view all the answers

What is discussed in Section 10-7?

The differences between prokaryotic and eukaryotic replication Signup and view all the answers

What is the function of the g-complex in DNA polymerase III?

To form a clamp that surrounds the DNA and slides along it during polymerization Signup and view all the answers

What is required for DNA polymerases to synthesize DNA?

A DNA template, a primer, and all four deoxyribonucleoside triphosphates Signup and view all the answers

What is the function of the 5' → 3' exonuclease activity of DNA polymerases?

To proofread and repair DNA during replication Signup and view all the answers

What is the primary function of DNA polymerase III?

To polymerize the newly formed DNA strand Signup and view all the answers

What is the function of the replisome in DNA replication?

To synthesize DNA on the leading and lagging strands Signup and view all the answers

What is the purpose of the RNA primer in DNA replication?

To provide a stable framework for the newly synthesized DNA strand Signup and view all the answers

What is the function of the 3' → 5' exonuclease activity of DNA polymerases?

To clear away short stretches of nucleotides during repair Signup and view all the answers

Why is the presence of a primer necessary for DNA polymerases to synthesize DNA?

Because DNA polymerases require a stable framework to start synthesizing DNA Signup and view all the answers

What is the main difference between the function of DNA polymerase I and DNA polymerase III?

DNA polymerase I is responsible for repair, while DNA polymerase III is responsible for replication Signup and view all the answers

What is the purpose of Mg2+ in DNA replication?

To facilitate the binding of DNA polymerases to the template Signup and view all the answers

What is the role of the 3'-hydroxyl group at the end of the growing chain?

It attacks the phosphorus adjacent to the sugar in the nucleotide to be added to the growing chain Signup and view all the answers

Which direction do all synthesis of nucleotides occur from the perspective of the growing chain?

5' to 3' Signup and view all the answers

What is the problem faced by the cell during DNA replication?

The two strands are going in opposite directions Signup and view all the answers

What is the name of the fragments formed during the synthesis of the lagging strand?

Okazaki fragments Signup and view all the answers

What is the role of DNA ligase?

To link the Okazaki fragments together Signup and view all the answers

How many DNA polymerases are present in E.coli?

At least five Signup and view all the answers

What is the function of polymerase II?

Repair Signup and view all the answers

What is the processivity of a DNA polymerase?

The number of nucleotides joined before the enzyme dissociates from the template Signup and view all the answers

What is the SOS response?

A mechanism of DNA repair Signup and view all the answers

What is the characteristic of polymerase I?

It is a single polypeptide chain Signup and view all the answers

What is the role of DNA gyrase in the conversion of relaxed, circular DNA to the supercoiled form?

To unwind the helix before sealing the nick Signup and view all the answers

What is the result of the replication fork moving in the absence of DNA gyrase?

The introduction of positive supercoils Signup and view all the answers

What is the function of the helicase protein in DNA replication?

To promote the unwinding of the DNA helix Signup and view all the answers

What is the role of the single-strand binding protein (SSB) in DNA replication?

To stabilize single-stranded regions of DNA Signup and view all the answers

What is the function of the primase enzyme in DNA replication?

To copy a short stretch of the DNA template strand to produce the RNA primer sequence Signup and view all the answers

What is the origin of the RNA primer in DNA replication?

The primase enzyme Signup and view all the answers

In which direction does DNA synthesis occur on both the leading and lagging strands?

From 5' to 3' Signup and view all the answers

What is the role of DNA polymerase III in DNA replication?

To synthesize new DNA strands Signup and view all the answers

What is the significance of the 3'-hydroxyl group on the RNA primer?

It serves as the attachment point for the growing DNA strand Signup and view all the answers

Why is RNA used as a primer in DNA replication?

Because it can be formed de novo without a primer Signup and view all the answers

What is the primary function of DNA polymerase a in eukaryotes?

To add primers to the template strands and synthesize a short nucleotide sequence Signup and view all the answers

Which DNA polymerase is involved in leading strand replication?

Polymerase e Signup and view all the answers

What is the role of PCNA in eukaryotic DNA replication?

To act as a sliding clamp Signup and view all the answers

What is the function of RFC in eukaryotic DNA replication?

To attach PCNA to polymerase d Signup and view all the answers

What is the role of FEN-1 and RNase H1 in eukaryotic DNA replication?

To remove the RNA primer Signup and view all the answers

What is the characteristic of DNA polymerase b?

It is a repair enzyme Signup and view all the answers

What is the characteristic of DNA polymerase g?

It carries out DNA replication in mitochondria Signup and view all the answers

What is the characteristic of DNA polymerases a and b in eukaryotes?

They lack exonuclease activity Signup and view all the answers

How do prokaryotes distinguish between the parental strand and the newly synthesized daughter strand during mismatch repair?

By the presence of methylated adenines Signup and view all the answers

Which enzyme is involved in the removal of the sugar and phosphate from the nucleotide during base-excision repair?

AP endonuclease Signup and view all the answers

What is the result of a double-stranded break (DSB) in DNA?

A threat to genome stability Signup and view all the answers

What is the term used to describe the exchange of one DNA sequence with another or the incorporation of a DNA sequence into another?

Genetic recombination Signup and view all the answers

What is the primary function of recombination in higher animals?

To create a diverse range of immunoglobulins Signup and view all the answers

What is the term used to describe areas of a chromosome with high recombination frequency?

Hot spots Signup and view all the answers

What is the name of the disease caused by defects in DNA repair mechanisms, resulting in numerous skin cancers at an early age?

Xeroderma pigmentosum Signup and view all the answers

Who demonstrated recombination using phages and bacteria?

Meselson and Weigle Signup and view all the answers

What is the role of MutH, MutS, and MutL in mismatch repair?

To form a loop between the mistake and a methylation site Signup and view all the answers

What is the process by which DNA strands exchange genetic material during recombination?

Strand invasion Signup and view all the answers

What is the name of the repair mechanism that involves the removal of a large section of DNA containing the lesion?

Nucleotide-excision repair Signup and view all the answers

What is the term used to describe abnormal chromosome numbers in gametes?

Aneuploidy Signup and view all the answers

What is the term used to describe the process of genetic recombination involving homologous sequences?

Homologous recombination Signup and view all the answers

What is the purpose of single-stranded binding proteins during mismatch repair?

To protect the template strand from degradation Signup and view all the answers

What is the protein product of the PRDM9 gene involved in?

Histone methyltransferase Signup and view all the answers

What is the name of the repair mechanism that involves the repair of DNA double-stranded breaks without a template?

Nonhomologous DNA end-joining Signup and view all the answers

What is a major difference between eukaryotic and prokaryotic DNA replication?

Number of origins of replication Signup and view all the answers

During which phase of the cell cycle does DNA replication occur?

S phase Signup and view all the answers

What is the purpose of the pathways that ensure DNA replication occurs only once per cell cycle?

To prevent errors in DNA replication Signup and view all the answers

Why is recombination critical during meiosis?

To ensure correct segregation of chromosomes Signup and view all the answers

What is the purpose of the origin recognition complex (ORC) in eukaryotic DNA replication?

To bind to the origin of replication and serve as an attachment site for other proteins Signup and view all the answers

What is the function of the replication activator protein (RAP) in eukaryotic DNA replication?

To activate the replication licensing factors (RLFs) Signup and view all the answers

What is the role of the replication licensing factors (RLFs) in eukaryotic DNA replication?

To make the DNA competent for replication Signup and view all the answers

What is the significance of the cyclin-dependent protein kinases (CDKs) in eukaryotic DNA replication?

They block the reassembly of the pre-replication complex (pre-RC) Signup and view all the answers

What is the purpose of the pre-replication complex (pre-RC) in eukaryotic DNA replication?

To license the replication of DNA Signup and view all the answers

How many different polymerases are present in eukaryotes?

19 Signup and view all the answers

What is the location of the polymerase g in eukaryotes?

Mitochondria Signup and view all the answers

What is the significance of the G1 phase in eukaryotic DNA replication?

It is the phase where the cell prepares for DNA replication Signup and view all the answers

What happens to the replication licensing factors (RLFs) after they are phosphorylated by the cyclin-dependent protein kinases (CDKs)?

They are released from the pre-replication complex (pre-RC) Signup and view all the answers

What is the role of the nuclear membrane in eukaryotic DNA replication?

It dissolves during mitosis, allowing the RLFs to enter the nucleus Signup and view all the answers

What is the primary function of DNA polymerase I in DNA replication?

Removing the RNA primer and replacing it with deoxynucleotides Signup and view all the answers

What is the family of ATPases that clamp loaders are members of?

AAA+ superfamily Signup and view all the answers

What is the function of the fingers domain in DNA polymerase?

Recognizing and binding to deoxynucleotides Signup and view all the answers

Which enzyme is responsible for sealing the nicks that remain after DNA synthesis?

DNA ligase Signup and view all the answers

How often do errors in replication occur spontaneously?

Once in every 10^4 to 10^5 base pairs Signup and view all the answers

What is the activity of DNA polymerase I that removes incorrect nucleotides?

3' exonuclease activity Signup and view all the answers

What is the role of DNA gyrase in DNA replication?

Introducing a swivel point in advance of the replication fork Signup and view all the answers

What is the result of the proofreading function of DNA polymerase I?

One error in every 10^9 to 10^10 base pairs Signup and view all the answers

What is the function of the clamp loader in DNA replication?

Opening the sliding clamp and inserting the DNA chain Signup and view all the answers

What is the process by which DNA polymerase I fills in behind it with its polymerase activity?

Nick translation Signup and view all the answers

What is the structure of the DNA polymerase?

A closed ring with domains referred to as fingers, palm, and thumb Signup and view all the answers

What is the function of the γ complex in E. coli?

Acting as a clamp loader Signup and view all the answers

What is the most common effect of ultraviolet light on DNA?

Creation of pyrimidine dimers Signup and view all the answers

What is the process by which enzymes recognize and repair mismatched base pairs?

Mismatch repair Signup and view all the answers

What is the direction of DNA synthesis on the lagging strand?

Discontinuous from 5' to 3' end Signup and view all the answers

What is the purpose of antioxidants in relation to DNA damage?

To prevent chemical damage caused by free radicals Signup and view all the answers

What is the role of primase in DNA replication?

Catalyzing the synthesis of an RNA primer Signup and view all the answers

What is the stoichiometry of Pol III enzymes in the replisome?

Three Pol III enzymes, with one for the leading strand and two for the lagging strand Signup and view all the answers

How do clamp loaders relate to DNA replication?

They are involved in sliding clamps Signup and view all the answers

What is the purpose of DNA polymerase I's 5' exonuclease activity?

To remove RNA primers Signup and view all the answers

Study Notes

Replication of DNA

DNA exists in many forms: single- and double-stranded, linear and circular
Many DNAs are double-stranded, and the process of replication is complex, with a high degree of fine-tuning to ensure fidelity
The cell faces three important challenges in replication:
- Separating the two DNA strands
- Synthesizing DNA from the 5' to the 3' end
- Guarding against errors in replication

Semiconservative Replication

DNA replication involves the separation of the two original strands and the production of two new strands with the original strands as templates
Each new DNA molecule contains one strand from the original DNA and one newly synthesized strand
This process is called semiconservative replication

How did scientists figure out that replication is semiconservative?

Experiments by Matthew Meselson and Franklin Stahl in the late 1950s used E. coli bacteria grown with a heavy isotope of nitrogen (15N)
The 15N-labeled DNA has a higher density than unlabeled DNA
The cells were then transferred to a medium with the usual isotope of nitrogen (14N), and the DNA was analyzed using density-gradient centrifugation
The results confirmed the predictions of semiconservative replication

Direction of Replication

Replication proceeds bidirectionally from the origin of replication
New polynucleotide chains are synthesized using each of the exposed strands as a template
Two replication forks advance in opposite directions from the origin of replication

DNA Polymerase

All synthesis of nucleotide chains occurs in the 5' to 3' direction
The last nucleotide added to a growing chain has a 3'-hydroxyl on the sugar
The incoming nucleotide has a 5'-triphosphate on its sugar
The 3'-hydroxyl group at the end of the growing chain is a nucleophile that attacks the phosphorus adjacent to the sugar in the nucleotide to be added

Semidiscontinuous DNA Replication

The two strands of DNA are going in opposite directions during replication
The problem is solved by different modes of polymerization for the two growing strands
One newly formed strand (the leading strand) is formed continuously from its 5' end to its 3' end
The other strand (the lagging strand) is formed semidiscontinuously in small fragments (Okazaki fragments)

Properties of DNA Polymerases

At least five DNA polymerases are present in E. coli
The properties of DNA polymerases include:
- Turnover number (speed of the synthetic reaction)
- Processivity (the number of nucleotides joined before the enzyme dissociates from the template)
- Proofreading and repair functions
DNA polymerase III has the highest turnover number and a huge processivity compared to polymerases I and II

Primase Reaction

RNA serves as a primer in DNA replication
The primer activity of RNA was first observed in vivo
A separate enzyme, called primase, is responsible for copying a short stretch of the DNA template strand to produce the RNA primer sequence
The primer and the protein molecules at the replication fork constitute the primosome

Synthesis and Linking of New DNA Strands

The synthesis of two new strands of DNA is begun by DNA polymerase III
The newly formed DNA is linked to the 3'-hydroxyl of the RNA primer
Synthesis proceeds from the 5' end to the 3' end on both the leading and the lagging strands
The RNA primer is removed by polymerase I, using its exonuclease activity
The primer is replaced by deoxynucleotides, also by DNA polymerase I
DNA ligase seals the remaining nicks

Architecture of DNA Polymerases and the Replisome

The various DNA polymerases have a common structure that is often compared to a right hand
The active site where the polymerase reaction is catalyzed lies in the crevice within the palm domain
The fingers domain acts in deoxynucleotide recognition and binding
The thumb is responsible for DNA binding

A Summary of DNA Replication in Prokaryotes

DNA synthesis is bidirectional
The direction of DNA synthesis is from the 5' end to the 3' end of the newly formed strand
One strand (the leading strand) is formed continuously, while the other strand (the lagging strand) is formed discontinuously
Five DNA polymerases have been found in E. coli
DNA gyrase introduces a swivel point in advance of the movement of the replication fork
A helix-destabilizing protein, a helicase, binds at the replication fork and promotes unwinding
The exposed single-stranded regions of the template DNA are stabilized by a DNA-binding protein
Primase catalyzes the synthesis of an RNA primer
The synthesis of new strands is catalyzed by Pol III
The primer is removed by Pol I, which also replaces the primer with deoxynucleotides
DNA ligase seals the remaining nicks### DNA Replication and Repair
DNA replication is an essential process that occurs only once in each cell generation, and it's crucial to ensure the fidelity of the replication process to prevent mutations.
The process of DNA replication is semiconservative, meaning that each new DNA molecule is composed of one old strand (the template) and one new strand synthesized during replication.

Proofreading and Repair

Proofreading refers to the removal of incorrect nucleotides immediately after they are added to the growing DNA during the replication process.
DNA polymerase I has three active sites, including the polymerase activity, proofreading activity, and 5' S 3' repair activity.
Proofreading improves the fidelity of replication to one error in every 10^9 to 10^10 base pairs.
The cut-and-patch process catalyzed by polymerase I takes place after polymerase III has produced the new polynucleotide chain.
Existing DNA can also be repaired by polymerase I using the cut-and-patch method.

Mismatch Repair

Mismatch repair is a process that recognizes and corrects errors in DNA replication.
Enzymes recognize incorrectly paired bases, and the area with the mismatch is removed, and DNA polymerases replicate the area again.
The challenge for the repair system is to know which of the two strands is the correct one.
Prokaryotes alter their DNA at certain locations by adding methyl groups, which helps the repair system distinguish between the parental and newly synthesized strands.

Nonhomologous DNA End-Joining (NHEJ)

NHEJ is a repair mechanism that handles double-stranded breaks (DSBs) in DNA.
A heterodimeric protein called Ku70/80 binds the broken ends of the DNA and recruits several other proteins that repair the damage.
This repair mechanism proceeds without a template, making it an error-prone mechanism.

Homologous Recombination

Homologous recombination is a natural process that rearranges genetic information to form new associations.
It involves the exchange of one DNA sequence with another or the incorporation of a DNA sequence into another.
The process is critical during meiosis, ensuring the correct segregation of chromosomes.
Hot spots are areas of a chromosome more likely to show recombination.

Eukaryotic DNA Replication

Eukaryotic replication is more complicated than prokaryotic replication, with multiple origins of replication, controlled timing, and more proteins involved.
Replication begins at multiple origins of replication, also called replicators, and occurs in the S phase of the cell cycle.
The process is tied to cell division, with the activation of cyclin-dependent protein kinases (CDKs) and the degradation of replication licensing factors (RLFs) to prevent reassembly of a pre-replication complex (pre-RC).

Eukaryotic DNA Polymerases

At least 19 different polymerases are present in eukaryotes, with 5 being studied more extensively.
The five best-studied polymerases are called a, b, g, d, and e, with different roles in DNA replication and repair.
Polymerase d is the principal DNA polymerase in eukaryotes, interacting with a protein called PCNA (proliferating cell nuclear antigen).

The Eukaryotic Replication Fork

The general features of DNA replication in eukaryotes are similar to those in prokaryotes.
The replication fork involves the formation of Okazaki fragments, initiated by Pol a, and the attachment of PCNA to Pol d.
The RNA primer is eventually degraded by separate enzymes, FEN-1 and RNase H1, and DNA ligase seals the nicks that separate the fragments.

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Description

Learn about the general features of DNA replication, including its process and forms, specifically in double-stranded DNA. This quiz covers the details of linear and circular DNA replication.