The new microscope opens up the molecular mysteries of cells

On the ' final boundary ' of science, the most difficult limit to overcome is to observe the molecular level activities in living cells. So far, a physicist at the University of Massachusetts Amherst has created a device to do this and began to uncover molecular level secrets like how enzymes regulate cellular functions.

Jennifer Ross has designed a kind of microscope that she calls the Single molecule TIRF (Single Molecule TIRF - the total internal reflection flourescence, the entire fluorescent reflection inside), which is much brighter than many commercial devices today. and has a great ability to observe and capture single molecules in real time.

The image of the katanin enzyme taken from the TIRF machine in Ross and colleagues' recent study is taken as a cover for the Biophysical magazine, along with their work showing that this achievement is the key to understanding the function of microtubules. - basic microtubule and harm in diseases related to their malfunction.

(Artwork: Popsci)

As the name suggests, these microtubules are always in good condition, hollow tubes about 25 nanometers in diameter to form bundles of fibers that form the structure for many cells in plants to humans. In plants, cellulose compounds are used to make plants strong. In humans, nerve axons cannot function properly without long and durable microtubule bundles that help extend their structure. Without these bundles, nerve cells shrink, causing neuromuscular diseases such as amyotrophic lateral sclerosis (ALS) or cerebral palsy. These microtubules play an important role in arranging materials inside cells in stages of mitosis and reducing feces.

Katanin is an enzyme that cuts microtubules in the middle or near the end, making it an important regulating factor in controlling these molecular structures. ' When you think of microtubules as a bundle of wooden columns you want to use to build houses, you have no way to cut them into reasonable lengths. You need katanin to cut it , 'Ross said.

However, in the long run it is not understood how this enzyme cuts microtubules. Another reason is that it is very difficult to clean them. Her postdoctoral assistant, Juan Daniel Diaz-Valencia, had a " remarkable effort " to purify katanin through a series of experiments at Umass Amherst in collaboration with David Sharp of Y Albert School Einstein. When Diaz-Valencia successfully cleaned katanin, ' he did it for two days continuously to get the data ,' she said.

In a recent series of experiments, her team not only documented katanin's cutting activity but also discovered that the activity depended on concentration. They proceeded to take katanin out of the cell and lead to the formation of microtubules that clog the inner cell area. ' Nobody has described it as simple as we do ,' Ross said. ' Because studying it in a large amount of solution is not effective. The Single Molecule TIRF microscope is necessary to observe exactly what happens. '

In the study, the scientists worked with purified pig brains, rich in microtubules, adding katanin with fluorescents to observe the mechanism of katanin cutting complexes taking place. Thanks to the TIRF microscope, the researchers filmed the video with a very sensitive camera that could see single light particles.

After a 3-minute control period from the start of the experiment to ensure that the microtubules do not self-transform, they have a control for each stage, the researchers add pure katanin to other concentrations. together. By measuring the brightness of fluorescent spots, they can count the number of molecules present more accurately than before, Ross said.

' What we found is that katanin was destroyed immediately and regenerated when used to cut the microtubules ,' said Professor. ' Now we know that katanin is a continuous regenerative subunit , it seems that you're constantly changing your car's steering wheel while you're driving .' But when the protein is breaking the microtubules, it also breaks and ends the experiment, so after 20 minutes the researcher has to repeat a new experiment.

Ross and his colleagues are taking the next step, designing a stronger microscope with high compatibility. They will focus their research on a relatively new enzyme, fidgetin, named after the experiment on a strain of mice with convulsions and jerky symptoms that push the head back and forth rather than nodding up and down as usual. These animals, noted for research in the 40s of the previous century, suffered a mutation in the production of fidgetin and caused this unusual seizure.

Just a decade ago, biologists began to test the function of fidgentin at the molecular level, Ross said. ' We found that this protein is very unique, it's very different from katanin and it regulates bone formation, ' she explains. ' Lack of this protein will cause muscle defects at birth. 'She and her colleagues are beginning to work with developmental biologists to design more in-depth experiments.

' We see unusual things like that, we can't help but learn it ,' she added. ' I will not be surprised if in the next 10 years we will be able to identify the bone defects of the newborn body caused by the lack of this protein .'