By creating a more real representation of the life of the DNA environment, researchers from the Northwestern University have discovered that the separation of threads, the essential process that is suffered by a double propeller “at rest” before it can start replication or make repairs, it can take more mechanical strength than the field believed previously.
Most biochemistry laboratories studying DNA isolate within a water -based solution that allows scientists to manipulate DNA without interacting with other molecules. They also tend to use heat to separate threads, heating the DNA to more than 150 degrees Fahrenheit, a temperature that a cell would never naturally reach. On the contrary, in a living cell DNA lives in a busy environment, and special proteins bind to DNA to mechanically relax the double propeller and then separate it.
“The interior of the cell is super full of molecules, and most biochemistry experiments are super unprocessed,” said Northwest professor John Marko. “You can think of extra molecules like billiard balls. They are hitting the double DNA propeller and preventing it from opening.”
Marko, professor of molecular biosciences, as well as physics at the Faculty of Weinberg Arts and Sciences of Northwestern, directed the research along with the postdoctoral researcher of the Northwest Parth Desai. In Marko’s laboratory, for his experiments, he and Desii use microscopic magnetic tweezers to separate the DNA and then carefully join the threads to the surfaces at one end, and small magnetic particles in the other, then perform high -tech images. Technology has existed for 25 years, and Marko was one of the first researchers who theorized and then used it.
Marko and Desai wrote the article that not only identifies, but quantifies the amount of stress imposed by overcrowding, which will be published on June 17 in the Biophysical Magazine
Desai introduced three types of molecules in the solution contained in the DNA to imitate proteins and investigated the interactions between glycerol, ethylene glycol and polyethylene glycol (each approximately the size of a double DNA propeller, two or three nanometers).
“We wanted to have a wide variety of molecules where some cause dehydration, destabilizing the DNA mechanically and then others that stabilize the DNA,” Desai said. “It is not exactly analogous to the things found in the cells, but it could be imagined that other competing proteins in the cells will have a similar effect. If they compete for water, for example, they would dehydrate the DNA, and if they do not compete for the water, they would be crowded with the DNA and would have this entropic effect.”
While it is fundamental, an investigation like this has been the basis of many, many, many medical advances, “said Marko, such as the deep sequencing of DNA, where scientists can now sequence an entire human genome in less than a day. He also believes that their findings can be widely applicable to other elements of fundamental biochemical processes.
“If this affects the separation of the DNA chain, all protein interactions with DNA will also be affected,” Marko said. “For example, the tendency that proteins adhere to specific sites in DNA and control specific processes; this will also be altered by overcrowding.”
In addition to executing more experiments that incorporate multiple overcrowding agents, the team hopes to approach a true representation of a cell and, from there, study how the interactions between enzymes and DNA are affected by overcrowding.
This work was supported by the National Health Institutes (subsidy R01-GM105847) and by subcontract to the 3D and Physics Center of the genome of the University of Massachusetts (Bajo NIH GRANT UM1-HG011536).