2024-04-12 18:07:14
Picture in your mind a traditional “landline” phone with a coiled wire connecting the receiver to the phone. The coiled telephone wire and the DNA double helix that stores the genetic material in every cell of the body have one thing in common; Both are convoluted, or twisted around themselves, and entangled in ways that can be difficult to undo. In the case of DNA, if this twisting is not dealt with, essential processes such as DNA copying and cell division stop. Fortunately, cells have an ingenious solution to carefully regulate DNA supercoiling.
In this study published in the journal knowledge, researchers at Baylor College of Medicine, Université de Strasbourg, Université Paris Cité and collaborating institutions reveal how DNA gyrase resolves DNA tangles. The findings not only provide new insights into this basic biological mechanism, but also have potential practical applications. Gyrases are biomedical targets for treating bacterial infections and the similar human versions of the enzymes are targets for many anticancer drugs. A better understanding of how gyrase works at the molecular level can improve clinical treatments.
Some DNA supercoiling is essential to make the DNA accessible to allow the cell to read and make copies of the genetic information, but too little or too much supercoiling is harmful. For example, the act of copying and reading the DNA throws it in front of the enzymes that read and copy the genetic code, interrupting the process. It has long been known that DNA gyrase plays a role in unwinding, but the details have been unclear.
DNA mini-circles and advanced imaging techniques reveal the first step to the DNA solution
We usually imagine DNA as the straight double helix structure, but inside cells, DNA exists in coiled loops. Understanding the molecular interactions between the supercoils and the enzymes involved in DNA functions has been technically challenging, so we typically use linear DNA molecules instead of coiled DNA to study the interactions. One of the goals of our lab was to study these interactions using a DNA structure that more closely mimics the actual twisted and looped form of DNA present in living cells.”
Dr. Lynn Zahidrich, study author, Kyle and Josephine Morro Chair in Molecular Virology and Microbiology and Professor in the Verna and March McLean Department of Biochemistry and Molecular Pharmacology at Baylor College of Medicine
After years of work, the Zechiedrich lab created small loops of coiled DNA. In fact, they took the familiar straight linear DNA double helix and twisted it in each direction once, twice, three times or more and joined the ends together to form a loop. Their previous research examining the three-dimensional structures of the coiled minicircles revealed that these loops form a variety of shapes that they hypothesized enzymes such as gyrase would recognize.
In the current study, their hypothesis was proven correct. The team of researchers combined their expertise to study the interactions of DNA gyrase with DNA minicircles using recent technological advances in electron cryomicroscopy, an imaging technique that produces high-resolution 3D views of large molecules, and other technologies.
“My lab has been interested for a long time in understanding how nanomolecular machines work in the cell. We studied the DNA gyrases, very large enzymes that regulate the winding of DNA,” said co-author Dr. Valerie Lamore, associate professor at the Institute of Genetics. et de Biologie Moléculaire et Cellulaire, Université de Strasbourg. “Among other functions, supercoiling is the cell’s way of confining about 2 meters (6.6 feet) of linear DNA into the cell’s microscopic nucleus.”
As the DNA coils inside the nucleus, it twists and folds into different shapes. Imagine twisting the telephone cord mentioned at the beginning, several times on itself. It will twist and form a loop by crossing DNA strands, tightening the structure.
“We found, just as we hypothesized, that gyrase is attracted to the mini-supercircle and positions itself in the interior of this coiled loop,” said co-author Dr. Jonathan Fogg, senior staff scientist of molecular virology and microbiology, and biochemistry and molecular pharmacology in the Zechidrich lab.
“This is the first step of the mechanism that drives the enzyme to resolve DNA tangles,” Lamore said.
“DNA gyrase, now surrounded by a tightly coiled loop, will cut one DNA helix in the loop, move the other DNA helix through the cut in the second, and reseal the break, which relaxes the twist and eases tangling, and regulates DNA coiling. Controlling the activity of the -DNA,” said Zachidrich. “Imagine you’re watching a rodeo. Like lassoing a cattle rope, superlooped DNA traps the gyrase in the first step. Gyrase then cuts one double helix of the DNA lasso and passes the other helix through the break to break free.”
Co-author Dr. Marc Nadal, professor at the Ecole Normale in Paris confirmed the observation of the looped DNA path around the gyrase using magnetic tweezers, a biophysical technique that allows measuring the distortion and fluctuations along the length of a single DNA molecule. Observation of a single molecule provides information that is often hidden when looking at thousands of molecules in traditional “ensemble” experiments in vitro.
It is interesting to note that the “DNA strand reversal model” for gyrase activity was proposed in 1979 by Dr. Patrick O. Brown and Nicholas R. The late Kozraly, also in a knowledge paper, long before researchers had access to coiled-coils or the enzyme’s three-dimensional molecular structure. “It’s especially meaningful to me that 45 years later, we’re finally providing experimental evidence to support their hypothesis because Nick was my postdoctoral mentor,” Zahidrich said.
“This work opens countless perspectives to explore the mechanism of this conserved class of enzymes, which are of great clinical value,” said Lamore.
“This work supports new ideas about how DNA activities are regulated. We propose that DNA is not a passive biomolecule activated by enzymes, but an active molecule that uses supercoils, loops and three-dimensional shapes to direct the accessibility of enzymes such as shears to sequences specific DNAs in a variety of situations, which will likely affect cellular responses to antibiotics or other treatments,” said Fogg.
Contributors to this work also include Marlène Vayssières (lead author), Nils Marechal, Long Yun, Brian Lopez Duran, and Naveen Kumar Murugasamy. The authors are affiliated with one or more of the following institutions: Baylor College of Medicine, Université de Strasbourg, Institut de Génétique et de Biologie Moléculaire et Cellulaire, INSERM, Université Paris and Hôpitaux Universitaires de Strasbourg.
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