Isolation of DNA Mutants

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Questions and Answers

What is the function of DnaB/helicase in bacterial chromosome replication?

To synthesize RNA primers

To bind to DnaA boxes and bend DNA

To unwind DNA in the 5' to 3' direction using energy from ATP (correct)

To methylate GATC sites

What is the role of DNA adenine methyltransferase (Dam) in regulating replication?

To initiate replication on fully methylated DNA

To unwind DNA at the origin of replication

To methylate the A on both strands of DNA (correct)

To synthesize RNA primers

What is the significance of the AT-rich region in the origin of replication (oriC)?

It is a region where strands separate during replication (correct)

It is a site for RNA primer synthesis

It is a site for DNA methyltransferase activity

It is a binding site for DnaA proteins

What is the function of DnaA boxes in the origin of replication (oriC)?

They are binding sites for DnaA proteins Signup and view all the answers

What is the result of the methylation of GATC sites?

Initiation of replication only occurs efficiently on fully methylated DNA Signup and view all the answers

What is the direction of DNA unwinding by DnaB/helicase?

5' to 3' Signup and view all the answers

What happens to the newly synthesized DNA strand immediately after replication?

It remains unmethylated Signup and view all the answers

What is the consequence of methylation of GATC sites on replication?

Initiation of replication only occurs efficiently on fully methylated DNA Signup and view all the answers

Which DNA polymerase is responsible for most of the DNA replication?

DNA pol III Signup and view all the answers

What is the function of the clamp loader complex in DNA polymerase III holoenzyme?

Helping the clamp protein bind to the DNA Signup and view all the answers

What is the function of the ε subunit in DNA polymerase III holoenzyme?

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

Which subunit of DNA polymerase III holoenzyme allows the enzyme to slide along the DNA without falling off?

β subunit Signup and view all the answers

Which DNA polymerase is involved in DNA repair and replication of damaged DNA?

DNA pol II, IV and V Signup and view all the answers

What is the function of DNA pol I?

Removes the RNA primers and replaces them with DNA Signup and view all the answers

How many subunits does DNA polymerase III holoenzyme consist of?

10 subunits Signup and view all the answers

What is the function of the α subunit in DNA polymerase III holoenzyme?

Synthesizes DNA Signup and view all the answers

What is the role of the β subunit in DNA polymerase III?

To form a dimer in the shape of a ring around template DNA, promoting the association of the holoenzyme with DNA Signup and view all the answers

Which of the following is released during the DNA polymerase-catalyzed reaction?

A pyrophosphate (PPi) Signup and view all the answers

What is the function of the helicase during DNA replication?

To unwind the double helix structure of DNA Signup and view all the answers

Which of the following genes is involved in the initiation of DNA replication?

DnaA Signup and view all the answers

What is the role of primase during DNA replication?

To synthesize the RNA primer Signup and view all the answers

What is the function of DNA polymerase III?

To catalyze the formation of a covalent bond between the incoming deoxyribonucleoside triphosphate and the 3'-OH of the previous deoxynucleotide Signup and view all the answers

Which of the following is a characteristic of DNA polymerase III?

It is a processive enzyme Signup and view all the answers

What is the result of the DNA polymerase-catalyzed reaction?

The formation of a covalent bond between the incoming deoxyribonucleoside triphosphate and the 3'-OH of the previous deoxynucleotide Signup and view all the answers

Study Notes

Isolation of Mutants

DNA mutants can be grouped into two categories when shifted to a non-permissive temperature:
- Those that show rapid arrest, indicating genes that encode enzymes necessary for DNA replication
- Those that complete their current round of replication but cannot start another, indicating genes that encode proteins necessary for initiation of replication
Table 11.3 summarizes the genes identified using this strategy, including mutations in DNA polymerase III, primase, helicase, DnaA, and DnaC

Chemistry and Accuracy of Bacterial DNA Replication

DNA polymerases catalyze the formation of a covalent bond between the incoming deoxyribonucleoside triphosphate and the 3'-OH of the previous deoxynucleotide
This process releases the last two phosphates of the incoming nucleotide as pyrophosphate (PPi)

DNA Polymerase III

DNA polymerase III is a processive enzyme, remaining attached to the template as it synthesizes the daughter strand
The β subunit of DNA pol III forms a dimer in the shape of a ring around template DNA, acting as a clamp protein
The clamp loader complex helps the clamp protein bind to the DNA

Bacterial Chromosome Replication

The process of bacterial chromosome replication is shown in Figure 11.18
An autoradiograph of E. coli in the act of replication is shown in Figure 11.19

Initiation of Replication

The origin of replication in E. coli is termed oriC
Three types of DNA sequences in oriC are functionally significant: AT-rich regions, DnaA boxes, and GATC methylation sites
Events that occur at oriC include:
- DnaA proteins binding to DnaA boxes and each other
- Additional proteins binding to bend DNA
- Strands separating at AT-rich regions
- DnaB/helicase unwinding DNA
GATC methylation sites regulate replication, ensuring only one round of replication occurs

DNA Polymerases

There are five DNA polymerases in E. coli: DNA pol I, II, III, IV, and V
DNA pol III is responsible for most DNA replication and is composed of 10 subunits
The α subunit catalyzes bond formation between adjacent nucleotides, while other subunits fulfill other functions
DNA pol I is composed of a single polypeptide and removes RNA primers and replaces them with DNA

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Description

This quiz is about the isolation of DNA mutants, specifically how they fell into two groups when shifted to a non-permissive temperature, and the genes encoded for DNA replication and initiation.