It may sound like science fiction, but inventors of a technique in which mixtures of biomolecules are frozen in time, blasted with an electron beam, and resolved into three-dimensional structures at atomic resolution just won this year’s Nobel Prize in chemistry. Still a work in progress, cryo-electron microscopy dramatically improved through decades of dedicated tinkering by Jacques Dubochet of the University of Lausanne in Switzerland, Joachim Frank of Columbia University in New York, and Richard Henderson of MRC Laboratory of Molecular Biology, Cambridge, England. The imaging feat has delivered a windfall of structures for biologists including neurodegenerative disease researchers, who in recent years got to lay eyes upon γ-secretase, Aβ fibrils, and tau fibrils at atomic resolution.
Inklings of cryo-EM arose in the 1970s, when Henderson and colleagues were attempting to discern the structure of bacteriorhodopsin. The protein refused to form crystals amenable to X-ray diffraction, so the researchers turned to electron microscopy instead. They blasted a frozen solution of the protein with a beam of electrons, which passed through the sample and were focused by a lens onto photographic film. That led to a three-dimensional structure of the protein in 1975, though its blob-like appearance was a far cry from the exquisite detail of today’s structures.
Over the following decades, Henderson, Frank, Dubochet, and a cadre of other researchers have improved every aspect of the technique. Dubochet developed a rapid freezing method using liquid ethane, which prevented biomolecules from drying out and distorting under the ensuing bombardment of electrons. Henderson and others created far more sensitive electron detectors, and Frank was instrumental in advancing computational algorithms that yielded a crisp, authoritative three-dimensional structure by merging millions of images.
All of these improvements have paid off. Rather than scratching their heads at the sight of the amorphous splotches of the early days, researchers now use cryo-EM to scrutinize relationships between individual amino acids within devilishly complex protein machines, such as the ribosome and spliceosome.
An atomic level structure of γ-secretase—the conglomerate that churns out Aβ—was a boon to Alzheimer’s researchers. An initial, near-atomic level structure revealed how each of the complex’s 20 transmembrane regions laid in the cell membrane, and assigned each to one of the four protein subunits (Sun et al., 2015). A few months later, an atomic-level structure unveiled interactions between specific amino acids, offering up clues about how mutations in the enzyme might lead to the overproduction of Aβ42 (Bai et al., 2015). Yigong Shi of Tsinghua University in Beijing led both studies.
Shi earned his cryo-EM chops with the help of Sjors Scheres—a cryo-EM expert Henderson recruited to the MRC. Scheres also developed an image-processing algorithm called RELION that helped take Shi’s initial γ-secretase structure to the atomic level. Scheres has put his stamp on other AD-related structures as well. This year, he collaborated with Michel Goedert, also at the MRC, to dazzle the field with atomic-level structures of tau fibrils (Fitzpatrick et al., 2017).
Scheres told Alzforum that amyloid proteins present a unique problem for cryo-electron microscopists. Compared with a protein complex like the ribosome, with its distinctive bumps and curves, amyloid fibrils are composed of meticulously stacked β-sheets, and thus are extremely smooth and difficult to get a handle on. “You have to start at the resolution where two β-strands can be separated,” he told Alzforum, and cryo-EM has finally advanced to the level where this is possible. Case in point, researchers led by Dieter Willbold and Gunnar Schröder at Heinrich-Heine-Universität, Düsseldorf, and the Forschungszentrum Jülich in Germany published an atomic-level structure of Aβ fibrils just last month using cryo-EM (Gremer et al., 2017).
As with most Nobel prizes, significant contributions in the field came from far more people than just the awardees. However, Scheres told Alzforum that he and others in the field largely agree that Henderson, Dubochet, and Frank made seminal advances deserving of the prize.
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Fitzpatrick AWP, Falcon B, He S, Murzin AG, Murshudov G, Garringer HJ, Crowther RA, Ghetti B, Goedert M, Scheres SHW. Cryo-EM structures of tau filaments from Alzheimer’s disease. Nature. 2017 Jul 13;547(7662):185-190. [PubMed].
Bai XC, Yan C, Yang G, Lu P, Ma D, Sun L, Zhou R, Scheres SHW, Shi Y. Nature. 2015 Sep 10;525(7568):212-217. An atomic structure of human γ-secretase. [PubMed].
Sun L, Zhao L, Yang G, Yan C, Zhou R, Zhou X, Xie T, Zhao Y, Wu S, Li X, Shi Y. Structural basis of human γ-secretase assembly. Proc Natl Acad Sci U S A. 2015 May 12;112(19):6003-8. [PubMed].
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Cheng Y, Grigorieff N, Penczek PA, Walz T. A primer to single-particle cryo-electron microscopy. Cell. 2015 Apr 23;161(3):438-449. [PubMed].
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