The genetic material inside every living cell, DNA, presents a significant packing challenge. If stretched out, the DNA from a single human cell would extend approximately 2 meters (6.5 feet). This immense length must be precisely organized and compacted to fit within the microscopic confines of a cell, such as a nucleus that is only about 10 micrometers wide. Cells employ a sophisticated coiling mechanism to manage DNA’s structure.
Defining Negative Supercoiling
DNA typically exists in a relaxed state known as B-DNA, where its two strands twist around a central axis approximately once every 10.4 to 10.5 base pairs. Supercoiling describes the additional twisting of the entire DNA double helix upon itself, similar to how a telephone cord might coil into larger loops. This coiling can be either positive or negative. Positive supercoiling occurs when DNA is overwound, adding more turns in the same direction as the natural helical twist, which tightens the molecule.
Negative supercoiling, in contrast, involves underwinding the DNA, meaning there are fewer twists per given length than in a relaxed state. This underwinding creates torsional strain, causing the DNA to coil back on itself in a left-handed direction, forming left-handed supercoils. The energy stored in negatively supercoiled DNA makes the double helix easier to separate, a property with significant biological implications.
The Biological Function of Negative Supercoiling
The torsional stress introduced by negative supercoiling is biologically advantageous, primarily because it facilitates the separation of the two DNA strands. This underwound state lowers the energy required to locally unwind the helix, making the DNA more accessible for cellular machinery. The stored energy in negative supercoils drives the transient unpairing of DNA bases, which is a prerequisite for two fundamental processes: DNA replication and transcription.
During DNA replication, the entire genome must be unwound and copied. Negative supercoiling at the origin of replication helps initiate this unwinding, allowing DNA helicases to separate the strands and DNA polymerases to begin synthesizing new DNA. Similarly, local unwinding of the DNA double helix is necessary to expose the genetic code for transcription. Negative supercoiling promotes this local strand separation, enabling RNA polymerase to bind to promoter regions and initiate RNA synthesis. Without this torsional stress, the energetic cost of unwinding DNA would be substantially higher, potentially slowing or halting these essential cellular activities.
Enzymatic Regulation of DNA Topology
The level of DNA supercoiling within a cell is tightly regulated by a family of enzymes known as topoisomerases. These enzymes manage DNA topology by introducing temporary breaks in the DNA strands, allowing the DNA to unwind or coil, and then resealing the breaks. Topoisomerases are categorized into two types based on their mechanism of action.
Type I Topoisomerases
Type I topoisomerases create transient single-strand breaks, allowing one DNA strand to pass through the other, primarily relaxing supercoils. For instance, bacterial topoisomerase I selectively relaxes negative supercoils.
DNA Gyrase (Type II Topoisomerase)
DNA gyrase, a type II topoisomerase predominantly found in bacteria, actively introduces negative supercoils into DNA. This process is energy-dependent, utilizing ATP hydrolysis to power the enzyme’s conformational changes. DNA gyrase works by binding to a DNA segment, wrapping approximately 130 base pairs around itself, and then creating a transient double-strand break. It then passes another segment of double-stranded DNA through this break before resealing the original break, changing the linking number by two and introducing two negative supercoils per step. This continuous activity ensures bacterial DNA remains negatively supercoiled, balancing positive supercoils that naturally arise ahead of moving replication and transcription machinery.
Therapeutic Targeting of Supercoiling Mechanisms
Understanding the mechanisms of DNA supercoiling and its enzymatic regulation has led to advancements in medicine, particularly in antibiotic development. Bacterial DNA gyrase is an appealing target for antimicrobial drugs because it is essential for bacterial survival and possesses structural differences from human topoisomerases. This allows for selective targeting, minimizing harm to human cells while disrupting bacterial processes.
The quinolone class of antibiotics, including drugs like ciprofloxacin, specifically targets bacterial DNA gyrase and topoisomerase IV. These drugs function by binding to the DNA-enzyme complex, stabilizing the transient double-strand breaks created by the topoisomerases. This stabilization prevents the resealing of DNA strands, leading to an accumulation of DNA breaks lethal to the bacterial cell. By interfering with the bacteria’s ability to manage their DNA topology, quinolone antibiotics halt DNA replication and transcription, leading to bacterial cell death.