It is a well-known fact that double-stranded DNA forms a double helix structure, but the double helix itself can twist and turn into different conformations. So, how does DNA change its shape, and why is this behavior important to our cellular functions? This video will use a physical model of DNA to help explain the significance of DNA topology and simulate how specific enzymes modify the conformation of double-stranded DNA. The unstrained, or relaxed, DNA structure has 10.4 base pairs per turn of the helix. One way to describe the structure of a DNA double helix is by a combination of its twist and writhe, the sum of which is called the linking number. The twist is described as the number of helical turns of one strand about the other and can be measured by counting how many times one strand wraps completely around the other. For example, this relaxed linear DNA has a twist of one, two, three. There is 0 writhe in this example. The writhe is described as how many times the double helix crosses over itself and is positive or negative depending on the orientation. For example, this circular DNA has a writhe of -1 because it follows the right hand rule and the upper strand travels left to right. And this DNA has a writhe of 1. Both of these examples also have twist. Realistically, long DNA molecules like these can have linking numbers in the thousands. For this video, we're going to look at a physical model of circular DNA with covalently linked ends. Unlike linear DNA, this shape is topologically constrained, meaning you can't change the linking number of the DNA without breaking one or both of the strands. But what would physically happen to this DNA if you added or removed twists or writhe in the double helix? Overwound DNA has fewer than 10.4 base pairs per turn of the helix. Thus, overwound DNA is more tightly wound than relaxed DNA and has a greater number of twists and a higher linking number than relaxed DNA. When DNA is overwound and has an increased linking number, the DNA becomes positively supercoiled, and the double-stranded structure begins wrapping around itself and creating positive writhe. Regardless of these visible changes in conformation, remember that positively supercoiled DNA will have a higher linking number than relaxed DNA, and the two strands of the double-stranded DNA are more difficult to separate. Underwound DNA has greater than 10.4 base pairs per turn of the helix. Thus, underwound DNA is less tightly wound than relaxed DNA and has a fewer number of twists and a lower linking number than relaxed DNA. When DNA is underwound and has a decreased linking number, the DNA becomes negatively supercoiled and the double-stranded structure begins wrapping around itself in the opposite direction as positive supercoiling, creating negative writhe. Regardless of these visible changes in conformation, remember that negatively supercoiled DNA will have a lower linking number than relaxed DNA, and the two strands of the double-strand DNA can be separated more easily. But if DNA is least strained in the relaxed form, how does the double helix change from being relaxed to overwound or underwound? And how does positively or negatively supercoiled DNA return to a relaxed state? Topoisomerases are a class of enzyme that usually restores DNA to its relaxed state. This means that topoisomerases unwind overwound DNA strands, reducing the linking number back to the preferred 10.4 base pairs per turn, or rewind underwound DNA strands, increasing the linking number back to 10.4 base pairs per turn. Topoisomerases work by breaking one or two DNA strands and passing the same number of DNA strands through the break. This change results in an increase or decrease in the linking number of our topologically constrained circular DNA. There are two main categories of topoisomerase, Type I and Type II. Type I topoisomerase breaks one of the two DNA strands and passes the other strand through the gap. This increases or decreases the linking number by 1. Here, the top DNA double helix represents the structure before the topoisomerase added a twist, and the bottom DNA double helix is after the enzyme completes the reaction. Type I topoisomerase reactions do not require additional energy. Type II topoisomerase breaks both of the two DNA strands and passes the entire double helix through the gap. This increases or decreases the linking number by 2. Here, the top DNA double helix represents the structure before the topoisomerase acts, and the bottom DNA double helix is after the enzyme completes the reaction. In this case, the result is that the writhe of the molecule increases by 2. To make this idea clear, this diagram shows what this change would look like if you added one positive writhe at a time instead of using Type II topoisomerase. Type II topoisomerase reactions require some form of energy, such as ATP or NADH. Bacteria also have a special form of Type II topoisomerases called gyrase. Gyrase enzymes use energy to negatively supercoil DNA rather than restoring DNA to the relaxed state. Certain thermophiles also have topoisomerases that use energy to positively supercoil DNA rather than restoring DNA to the relaxed state. Because it acts oppositely to gyrase, these enzymes are called reverse gyrase. But if DNA is unstrained in its relaxed state, why would organisms have enzymes that intentionally supercoil their DNA either positively or negatively? Thermophiles and hyperthermophiles thrive at relatively high temperatures, ranging from 45 to over 100 degrees Celsius. Positive supercoiling in thermophile DNA adds additional twists or writhe to the double helix, thus increasing the linking number. These changes stabilize the structure, preventing the DNA from denaturing as easily at high temperature. Wanting stable DNA structures makes evolutionary sense, but what about the organisms with negatively supercoiled DNA? In fact, the DNA of most organisms is negatively supercoiled. Negatively supercoiled DNA provides a store of free energy that helps with cellular processes that require strand separation of the double helix, like DNA replication and transcription. Underwound DNA has a tendency to partially separate, so strand separation is easier than in a relaxed DNA structure, with more twists holding the double helix together. In short, negatively supercoiled, or underwound, DNA makes it easier to separate the double helix into two single strands. As you separate the double helix and negatively supercoiled DNA, you are creating more twists in the rest of the DNA, causing rewinding of the underwound strands. Rather than putting strain on the DNA topology, the DNA that is still base paired is returned to the ideal relaxed state. Plus, you have a portion of separated single DNA strands that can be used in cellular processes like replication and transcription. If you tried to separate the double helix of relaxed DNA, you would introduce more twists in the DNA and end up overwinding, or positively supercoiling, the double helix, which is energetically unfavorable. This video modeled how DNA topology extends beyond the double helix structure and how topology is critical to our cell's ability to function smoothly by replicating or transcribing DNA more efficiently. Now, are you able to model overwound and underwound DNA and tell the difference between Type I and Type II topoisomerases and their functions? And can you explain what role DNA topology plays in cellular processes and function? Thanks for watching.