Question

Draw a mechanism for the topoisomerase-catalyzed cleavage of DNA that gives the free \(5^{\prime}\)-end.

Short answer

The mechanism involves the enzymatic action of topoisomerase causing a break in the DNA's phosphate backbone. As a result, the strands separate, resulting in the emergence of a free 5'-end.

Step-by-step solution

Understanding Enzymatic Action of Topoisomerase

Topoisomerase is an enzyme that modulates the overwinding or underwinding of DNA. The enzyme breaks the DNA's phosphate backbone to facilitate the unwinding process.

Identifying the Formation of Free 5'-End

When DNA is broken by topoisomerase, it results in the release of free ends. In this case, we focus on the release of the 5'-end. This end usually has a phosphate group attached.

Drawing the Mechanism

For simplicity, represent DNA as a double helix structure, and topoisomerase as an entity interacting with the DNA molecule. At the cleavage site, illustrate the breaking of the DNA backbone, leading to the separation of DNA strands and the free 5'-end emerged.

Highlighting the Resultant 5'-End

In the final step, draw attention to the free 5'-end by marking it. This is the result of topoisomerase action.

Key concepts

Topoisomerase Mechanism

Topoisomerases are crucial enzymes that manage the topological states of DNA. They are responsible for resolving DNA supercoiling, a condition that arises when DNA is overwound or underwound. During replication, transcription, or other cellular processes, the DNA has to struggle against tensions caused by its coiled nature. Topoisomerases temporarily break the DNA's phosphate backbone, allowing it to unwind or rewind to resolve these tensions.

• Type I Topoisomerases - These enzymes cut one strand of the DNA helix, allowing the other strand to rotate around it, which relieves the tension. Then, the cut strand is ligated back together.
• Type II Topoisomerases - These make transient double-strand breaks, enabling one segment of DNA to pass through another. These enzymes are vital during processes like chromosomal segregation.

The topoisomerase mechanism involves a fascinating chemical process where the enzyme orchestrates the cleavage and re-ligation of DNA strands, ensuring its integrity and functionality.

DNA Cleavage

The process of DNA cleavage by enzymes like topoisomerases is pivotal for cellular DNA management. Cleavage refers to the breaking of the phosphodiester bond in the DNA backbone. This act is strategic, allowing for temporary adjustments in the DNA's form without altering its sequence.
When topoisomerase cleaves the DNA, it does so in a controlled manner using a catalytic tyrosine residue. This residue attacks the DNA strand phosphate, forming a covalent bond and leading to a temporary break. Post-cleavage, the DNA can be manipulated, such as unwinding to relieve supercoils, before resealing the strand.
This precision cut helps to maintain genome stability and thus, the proper functioning of the cell.

5'-end DNA Structure

When discussing DNA ends, there are two critical configurations: the 5'-end and the 3'-end. These notations reflect the carbon numbering in DNA's sugar phosphate backbone. The 5'-end holds a terminal phosphate group, which plays a crucial role when DNA is cleaved by topoisomerases.
During topoisomerase action, the cleavage creates a free 5'-end. This creates a single-stranded break in the DNA, with a phosphate residue still bonded to the 5' carbon of deoxyribose. This configuration ensures the DNA's ability to be swiftly ligated back after the topological problem is solved.

• The free 5'-end is essential for DNA template elongation during replication
• Crucial for joining Okazaki fragments on the lagging strand during DNA synthesis

Understanding the nature of the 5'-end and its interactions within the cell is key to comprehending DNA repair and replication dynamics.