Biology Chapter 5: Topoisomerases and Nucleases

Questions and Answers

What is the primary function of Type II topoisomerase in relation to helical turns of DNA?

It changes the DNA by two helical turns. (correct)

It fixes single-stranded breaks in DNA.

It introduces positive supercoils.

It unwinds DNA completely without cleaving.

What effect does negatively supercoiled DNA have on cellular processes?

It prevents transcription initiation.

It inhibits DNA replication.

It promotes strand separation. (correct)

It facilitates DNA denaturation.

Which of the following describes the end result of Type II topoisomerase cleavage and passage?

It religates both strands of the cleaved segment. (correct)

It forms a single-stranded break in DNA.

It prevents DNA tangling.

It creates two separate DNA molecules.

What is the structure of cellular DNA typically considered?

Negatively supercoiled. Signup and view all the answers

How does Type II topoisomerase achieve the passage of DNA strands?

Via a double-stranded break. Signup and view all the answers

Why is positively supercoiled DNA advantageous for thermophilic organisms?

It resists heat denaturation. Signup and view all the answers

What role does reverse gyrase play in thermophilic organisms?

It introduces positive supercoils. Signup and view all the answers

What happens to DNA during the bind-cleavage (double)-passage-religation mechanism of Type II topoisomerase?

Double-stranded breaks and strand passage occur. Signup and view all the answers

What effect do topoisomerases have on DNA supercoiling?

They can increase or decrease supercoiling. Signup and view all the answers

Which of the following best describes the function of Type I topoisomerase?

It creates a single-stranded break to relax supercoils. Signup and view all the answers

What is produced by each round of catalysis in Type I topoisomerase activity?

One helical turn change, relaxing one negative supercoil. Signup and view all the answers

What is the first step in the Type I topoisomerase mechanism?

Binds to a segment of double-stranded DNA. Signup and view all the answers

How does the Type I topoisomerase mechanism change DNA's structure?

It relaxes negative supercoils by changing DNA's helical turns. Signup and view all the answers

What role does the cleaved DNA strand play during the activity of Type I topoisomerase?

It passes through the break formed by the enzyme. Signup and view all the answers

Which type of topoisomerase is characterized by cutting both strands of DNA?

Type II topoisomerase. Signup and view all the answers

What happens during the 'reverse reaction' in Type I topoisomerase?

The cleaved strand is re-ligated, restoring DNA integrity. Signup and view all the answers

What happens to circular relaxed DNA when it is subjected to positive supercoiling?

It forms more helical turns. Signup and view all the answers

What role does topoisomerase I play in the context of DNA supercoiling?

It relaxes supercoiled DNA. Signup and view all the answers

In an agarose gel electrophoresis, how do DNA bands typically migrate?

From top to bottom. Signup and view all the answers

What is the visual marker used to stain DNA in agarose gel electrophoresis?

Ethidium bromide. Signup and view all the answers

What condition results in the formation of a negative supercoil in DNA?

If it has fewer helical turns. Signup and view all the answers

Which type of supercoil is produced when circular relaxed DNA is underwound?

Negative supercoil. Signup and view all the answers

What is the configuration of circular relaxed DNA in terms of helical turns?

It has one helical turn per 10.5 bp. Signup and view all the answers

What would be the effect of applying topoisomerase I to supercoiled DNA?

It would convert it to relaxed DNA. Signup and view all the answers

What is the product of RNA 5’ pApGpCpUpCpGpUpC 3’ after being digested with RNase T1?

pApGp + CpUpCpGp + UpC Signup and view all the answers

Which statement accurately reflects the action of DNase I on RNA-DNA fusion molecules?

It produces RNA fragments and non-selectively cleaves DNA. Signup and view all the answers

What distinguishes RNase A from RNase T1?

RNase A is specific for pyrimidines. Signup and view all the answers

What is a characteristic of alkaline hydrolysis of RNA?

It requires the presence of 2’-OH to act on RNA. Signup and view all the answers

What is the end product of RNA after its hydrolysis by concentrated NaOH?

RNA is completely hydrolyzed into nucleotides. Signup and view all the answers

What occurs when DNA is supercoiled?

The DNA strands partially relax. Signup and view all the answers

Which enzyme specifically degrades RNA?

Ribonuclease Signup and view all the answers

In the context of nucleases, what is the difference between an endonuclease and an exonuclease?

Endonucleases hydrolyze interior phosphodiester bonds. Signup and view all the answers

What characteristic does negatively supercoiled DNA exhibit in gel migration?

It migrates slower as it is less compact. Signup and view all the answers

Which statement about the nature of phosphodiester bond cleavage is true?

It produces both 5’-OH, 3’-P and 3’-OH, 5’-P ends based on the enzyme. Signup and view all the answers

How is the supercoiling state of DNA quantified?

Using the formula S = Lk - Lko. Signup and view all the answers

What is a key feature of a nuclease that exhibits substrate specificity?

It requires a specific type of nucleic acid to function. Signup and view all the answers

What type of cleavage does DNase I produce?

Only 3’-OH and 5’-P ends. Signup and view all the answers

Study Notes

Topoisomerases

Topoisomerases alter DNA supercoiling; classified into type I and type II.
Type I topoisomerases relax DNA by cutting one strand (creating a single-stranded break) and allowing the other strand to pass through before re-ligating the cut strand.
Each action of type I topoisomerase results in the relaxation of one negative supercoil.

Mechanism of Type I Topoisomerases

Binds to double-stranded DNA, cleaves, passes the unbroken strand through the break, and then religates the cut strand.
Each cycle changes the DNA by one helical turn, reducing supercoiling effectively.

Type II Topoisomerases (DNA Gyrase)

Type II topoisomerases bind to two segments of DNA, cleave both strands at one segment (creating a double-stranded break), pass another segment through the break, and then religate both strands.
Catalysis results in changes of two helical turns, generating two negative supercoils and aiding in untangling DNA.

DNA Supercoiling

Cellular DNA is typically negatively supercoiled, promoting easier strand separation which is necessary for replication and transcription.
Thermophilic organisms contain positively supercoiled DNA, providing structural stability at high temperatures. Reverse gyrase generates positive supercoils in these organisms.

Experimental Analysis of Supercoiling

Circular DNA supercoiling can be visualized using agarose gel electrophoresis; relaxed DNA moves differently than supercoiled DNA.
Supercoiled DNA treated with topoisomerase I relaxes to a single supercoil, observable on a gel.

Nucleases

Nucleases catalyze hydrolysis of phosphodiester bonds in nucleic acids, aiding in DNA and RNA degradation processes.
Types include ribonuclease (RNase) degrading RNA, deoxyribonuclease (DNase) degrading DNA, and non-specific nucleases acting on both.
Nucleases vary by position of attack: endonucleases cleave bonds within the chain, while exonucleases cleave from chain ends.

Phosphodiester Bond Reactions

RNases and DNases produce 5'-OH and 3'-P or vice versa, depending on the specific nuclease.
Specificity can vary; some nucleases target specific bases while others can act more broadly.

Alkaline Hydrolysis of RNA

Concentrated NaOH can hydrolyze RNA but does not affect DNA due to DNA's lack of a 2'-OH group, which prevents similar reactions.
Alkaline treatments lead to RNA denaturation by breaking hydrogen bonds without damaging DNA structurally.

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Explore the functions and classifications of topoisomerases in DNA supercoiling through this quiz. Understand the differences between Type I and Type II topoisomerases and their roles in DNA manipulation. Test your knowledge on these essential enzymes crucial for DNA structure and function.