The story of cryo-EM started when Richard Henderson used electron microscopy to determine a three-dimensional model of bacteriorhodopsin by averaging multiple images obtained with weak electron beams. Bacteriorhodopsin is a protein, present in cell membrane of bacteria. Its function is to pump hydrogen ions across cell membranes. He observed that when electron beams are put on a biological sample there is radiation damage that destroys the structure. In all the early applications of EM on live sample, the samples were destroyed. Jacques Dubochet found a solution to the problem. He studied the behaviour of water which is the environment in which all the bio molecules are present. On freezing water, it forms ice. Ice can be a hexagonal crystal or cubic one. But if you freeze water very rapidly rather than forming a crystal structure (like salt or diamond), it freezes into an amorphous structure (like talcum powder). This process of rapid freezing of water is called vitrification. Cooling the sample reduces the effect of radiation damage by a factor of 4 to 5. He used liquid ethane at -196C for this purpose. At this temperature along with reduction in radiation damage, the sample is also protected from dehydration in the vacuum chamber of electron microscope. It allows the bio-molecules to retain their shape in a vacuum. But to obtain a complete 3D structure of a protein one 2D image is not enough. A series of 2D images are taken at different angles which are analyzed by computer software to reconstruct accurate and detailed 3D models of intricate biological structures at sub-cellular and molecular scales. This algorithm was made by Joachim Frank in 1975. These models can show interactions that were impossible to visualize previously, a key to scientific experiments. In 1990, Richard Henderson was the first to use a cryo-electron microscope to generate a 3D image of a protein at atomic resolution. He demonstrated that it is possible to obtain atomic resolution structures of bio-molecules using cryo-EM.
Electron cryo-microscopy has become an important and a very dominant method in structural biology. Structures obtained by cryo-EM and related techniques are of fundamental importance for understanding life chemistry, and they can help scientists develop drugs by elucidating the way bioactive agents interact with bio-molecules. During the recent Zika virus outbreak, cryo-EM was used to obtain the structure of Zika virus. It helped to understand how the virus infects humans and causes different symptoms. This knowledge was used to develop protocols to combat the outbreak and develop vaccines against the virus.
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2. B Stu, Cryo-electron microscopy innovators win 2017 Nobel Prize in Chemistry. Cemical and engineering news, ACS, 04-10-2017
3. Royal Swedish academy, Scientific Background: The development of cryo-electron microscopy. https://www.nobelprize.org/nobel_prizes/chemistry/laureates/2017/advanced.html