The Fantastic Cryogenic electron microscopy

Posted by beauty33 on March 15th, 2019

What is a Cryogenic electron microscopy (Cryo-EM)? Why do we call it “fantastic”?

First of all, the cryo-electron microscope is a kind of electron microscope. EM is an abbreviation for electron microscopy. The so-called electron microscope is developed on the basis of optical microscope. Optical microscopy uses visible light as a probe to observe microscopic objects, such as optical microscopy. However, for protein molecules in cells, it is invisible to the optical microscope.

Why is that? The reason is actually very simple. Optical microscopy uses the volatility of photons, and the wavelength of photons is about 500 nm. The size of the protein molecule is between 1-100nm, so the wavelength of the photon is larger than that of the protein molecule. Therefore, the light on the protein molecule is like a duck going to "step on" an ant, which seems to be impossible. Light waves can go through protein molecules. So, you can't see proteins via an optical microscopy. In order to "step on" an ant, there must be a needle smaller than the ant. This is the source of electron microscopy. The wavelength of an electron is about one of the hundred thousandth of the wavelength of a photon, meaning that it’s a very fine probe. Theoretically, it can be reflected on biological macromolecules such as protein molecules. These reflected electrons can produce a photo. This is the basic principle of electron microscopy.

However, electron microscopy is only theoretically able to see protein molecules. Because electron microscopy can only be used to observe some inorganic samples. For example, it can be used to observe the surface of graphite and ceramics. But once the electrons of the electron microscope are bombarded with biological macromolecules such as protein molecules, the problem arises.

The first problem is about vacuum. Electrons can only maintain stable kinetic energy when flying in a vacuum. The biological macromolecules such as proteins are generally in solutions, and in a vacuum environment, the solution will evaporate and contaminate the electron microscope.

The second problem is that electrons are likely to break down proteins on biological macromolecules such as proteins because the energy of electrons are relatively high, and biomacromolecules generally rely on hydrogen bonds to maintain their spatial structure. The energy of hydrogen bonds is very low. So, after the electrons were shot to the hydrogen bonds, they will be destroyed.

The third problem is even more serious, because biological macromolecules such as protein molecules are active, they are moving, and the direction in which electrons are reflected back will become disorganized by the movement of molecules. It's like we're shooting a racing game with a normal camera, and the photos we take are all fuzzy—the reason is simple: the car is moving.

In order to observe the spatial structure of biological macromolecules such as protein structure and to study the relationship between spatial structure and their functions, scientists need to immobilize protein molecules in a vacuum environment. Therefore, scientists put forward the method of freezing. They quickly freeze biological samples containing water with liquid ethane, etc., so that a very thin water film can be prepared (biomolecules are frozen in this water film). After the freezing is completed, electron microscopy can be used to observe the spatial structure of biological macromolecules such as proteins. In order to resolve the relationship between many of the captured two-dimensional photos and the three-dimensional structure of biomacromolecules, a set of relevant computer algorithms is needed.