Cryoelectron Microscopy - Cryo-EM
Cryo-electron microscopy (cryo-EM: cryogenic electron microscopy) is an imaging method based on the elastic scattering of a beam of electrons passing through thin layers of studied materials or molecules suspended in ice. Electrons scattered during transmission create its two-dimensional, enlarged projection on the detector. Due to the very short wavelength of electrons, it is possible to achieve atomic resolution of the measurement.
In order to perform cryomicroscopic measurements of proteins or other biomolecules, the suspended particles with the sample are frozen in liquid ethane. Thanks to this, unlike crystallography, it is possible to visualize moving, heterogeneous objects and to study the conformational changes of a given molecule. Their three-dimensional structure is reconstructed on the basis of automatic analysis of hundreds of microscopic images of single molecules, showing their two-dimensional projections in different, random orientations. Contrary to X-ray crystallography, this method does not require prior crystallization of biomolecules.
The Cryo-EM method is widely used in the pharmaceutical and biotechnology industries, and its creators, Jacques Dubochet, Joachim Frank, and Richard Henderson, were awarded the 2017 Nobel Prize. The remarkable usefulness of this technique was confirmed in March 2020, when scientists managed to solve at an unprecedented rate the structure of the SARS Cov2 coronavirus spike protein https://science.sciencemag.org/content/367/6483/1260, which made it easier for scientists and doctors to fight the COVID-19 pandemic
Who can be interested in Cryo-EM?
For science and industry. The applications of cryo-EM are focused on medical and pharmaceutical biotechnology, broadly understood experimental biology - biochemistry, molecular biology, cell biology and biophysics. One of the main applications is structural biology, and cryo-EM studies have made it possible to understand, among other things, the processes of reading genetic information or conducting signals in the nervous system. An example is the detection of protein build-up in patients with Alzheimer's disease. An impressive achievement in recent months is the precise determination of the chemical structure of such deposits, isolated directly from the patient's brain. The above discoveries are important not only for understanding the mechanisms of disease formation but also for developing new effective therapies.
The microscope can be used to study polymer compounds, semiconductor materials, image cell organelles, assess the structure and dynamics of membrane proteins, three-dimensional imaging the molecular structure of viruses, as well as analyze the stages of infecting cells with viruses and their reproduction, in the analysis of the antibacterial activity of bacteriophages in phage therapy in infections antibiotic-resistant strains, analysis of microbubble dynamics, telomere structure in the context of anti-cancer therapy and molecular aspects of longevity, analysis of drug transport across membranes, antibody-virus and bacterial interaction, the structure of parasitic protozoa and their interactions with tissues, as well as many other research topics.
For the pharmaceutical industry. The "resolution revolution" with cryo-EM microscopes allows for a resolution of <2Å, capable of establishing a structure-activity relationship. In particular, companies use cryo-microscopes to understand the specific binding mode of drug molecules and to study the surrounding space in the binding pockets to further fine-tune specificity and increase their affinity. In addition, this technology can be used to design and understand the shape of the drug molecule itself (e.g., antibodies and aptamers), even in the absence of a target protein. The idea of incorporating cryomicroscopy into drug development has emerged from the recently formed Cambridge Pharma EM consortium that connects Cryo-EM microscopes in an academia (MRC LMB) with five major pharmaceutical companies (Astex, AstraZeneca, GSK, Heptares and UCB).