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Proc Natl Acad Sci U S A. 2020 Jan 14;117(2):1009-1014. doi: 10.1073/pnas.1919114117. Epub 2019 Dec 26.

High-yield monolayer graphene grids for near-atomic resolution cryoelectron microscopy.

Author information

1
Department of Molecular Biology, Princeton University, Princeton, NJ 08544; yimoh@princeton.edu nyan@princeton.edu.
2
Department of Molecular Biology, Princeton University, Princeton, NJ 08544.
3
Department of Electrical Engineering and Computer Science, Massachusetts Institute of Technology, Cambridge, MA 02139.
4
Department of Physics, Princeton University, Princeton, NJ 08544.
5
PRISM Imaging and Analysis Center, Princeton University, Princeton, NJ 08544.

Abstract

Cryogenic electron microscopy (cryo-EM) has become one of the most powerful techniques to reveal the atomic structures and working mechanisms of biological macromolecules. New designs of the cryo-EM grids-aimed at preserving thin, uniform vitrified ice and improving protein adsorption-have been considered a promising approach to achieving higher resolution with the minimal amount of materials and data. Here, we describe a method for preparing graphene cryo-EM grids with up to 99% monolayer graphene coverage that allows for more than 70% grid squares for effective data acquisition with improved image quality and protein density. Using our graphene grids, we have achieved 2.6-Å resolution for streptavidin, with a molecular weight of 52 kDa, from 11,000 particles. Our graphene grids increase the density of examined soluble, membrane, and lipoproteins by at least 5-fold, affording the opportunity for structural investigation of challenging proteins which cannot be produced in large quantity. In addition, our method employs only simple tools that most structural biology laboratories can access. Moreover, this approach supports customized grid designs targeting specific proteins, owing to its broad compatibility with a variety of nanomaterials.

KEYWORDS:

UV/ozone; cryo-EM; graphene grid; high resolution; structure determination

PMID:
31879346
PMCID:
PMC6969529
[Available on 2020-06-26]
DOI:
10.1073/pnas.1919114117

Conflict of interest statement

Competing interest statement: The authors N. Yan and Y.H. are inventors on the patent application entitled “Fabrication of nanomaterial cryogenic electron microscopy (cryo-EM) grids.”

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