A classical lay summary by Emery Haley, Van Andel Institute. 

Knowing the structure of biological molecules is important because it improves our understanding of the function of the molecule. It can also be valuable for guiding the design of new drugs. The same way a locksmith needs to know where the tumblers are placed inside of a lock before designing a perfectly fitting key, structural information allows scientists to design drugs that can interact specifically with the intended molecule. This specific interaction increases effectiveness and reduces side-effects.

Historically, humans have been limited to understanding structural details that we could see with just our eyes, then with a magnifying lens, and eventually with a light microscope. To see details even smaller than the wavelength of visible light, we need to use X-rays or a beam of electrons. X-ray diffraction patterns were first used to determine the structure of table salt crystals in 1913 (Bragg 1913). Then the technique was adapted to determine the structures of organic compounds and biological molecules. It was famously used by Rosalind Franklin (Franklin and Gosling 1953) to help determine the structure of the DNA double helix.

Since their invention in the 1930s, electron microscopes have been used to examine the structure of inorganic molecules, but biological molecules are extremely sensitive to damage from the electron beam. Similar to X-ray crystallography, early electron microscopy of biological molecules was performed by imaging a crystal of the molecule (Henderson 1995). Unfortunately, most biological molecules are extremely difficult or impossible to crystallize for imaging. The crystal state can also be very different from the natural state of the molecule in a cell.

In the 1970s, Joachim Frank discovered a way to compute 3D structures from individual molecules in solution, rather than from crystals (Frank 1975). Because the molecules are so tiny, each individual image is unclear, like when you try to take a picture with a camera phone using the maximum zoom. By taking millions of images of the molecule and using a computer to line them all up and average them together, we can get a clear image. Combining these clear averaged images from many different angles then creates a 3D view of the structure.

Finally, Jacques Dubochet determined a way to freeze the molecules for imaging (Dubochet et al. 1988). His method, called “vitrification”, flash freezes a thin layer of the molecules in solution, preserving the shape of the suspended molecule. This water-like ice is called “vitreous ice”. Unlike like regular crystal ice, it does not interfere with the electron beam. This “vitreous ice” also protects the molecules from damage by the electron beam.

The 2017 Nobel Prize in Chemistry was awarded to Jacques Dubochet, Joachim Frank, and Richard Henderson for their developments that made it possible to determine structures of biological molecules in solution using cryo-electron microscopy (Cryo-EM). Since 2012, further advances in electron detection and image analysis have led to the “resolution revolution” and an explosion in the availability of high-resolution structures of protein and other complex biological molecules.

Graphic: https://people.csail.mit.edu/gdp/cryoem.html

Bragg, W.L. 1913. The structure of some crystals as indicated by their diffraction of X-rays. Proceedings of the Royal Society of London. Series A, Containing papers of a mathematical and physical character 89(610): 248-277.

Dubochet, J., M. Adrian, J.-J. Chang, J.-C. Homo, J. Lepault, A.W. McDowall, and P. Schultz. 1988. Cryo-electron microscopy of vitrified specimens. Quarterly reviews of biophysics 21(2): 129-228.

Frank, J. 1975. Averaging of low exposure electron micrographs of non-periodic objects. In Single-Particle Cryo-Electron Microscopy: The Path Toward Atomic Resolution: Selected Papers of Joachim Frank with Commentaries, World Scientific, pp. 69-72.

Franklin, R., and R. Gosling. 1953. The structure of sodium thymonucleate® bers. I. The influence of water content. Acta Crystallog 6: 673-677.

Henderson, R. 1995. The potential and limitations of neutrons, electrons and X-rays for atomic resolution microscopy of unstained biological molecules. Quarterly reviews of biophysics 28(2): 171-193.