Visualizing macromolecular structures in situ by cryoEM and cryoET

Session

Public Health: The Impact of Microscopy

Authors

Prof Peijun Zhang (1)

Affiliations

1. University of Oxford

Keywords

in situ, structural biology, cryoEM, cryoET, subtomogram averaging, viruses

Abstract text

Cryo-electron microscopy (cryoEM) is a powerful method for the high-resolution three-dimensional structural characterization of a wide range of biological samples in a close-to-native, frozen-hydrated state. Such biological samples, preserved in vitrified ice, are extremely radiation sensitive, therefore images of these have low signal-to-noise ratios and low contrast [1]. Recent development in microscope instrumentation, direct electron detector, microscope automation and high throughput imaging, and advanced software for data processing and image reconstruction, has revolutionized the field of structural biology, allowing protein structures to be determined at the atomic resolution, especially using cryoEM SPA method (Fig. 1). For studying macromolecular complexes that are intrinsically flexible and dynamic, and often function in higher-order assemblies that are difficult to purify, cryoET and subtomogram averaging (cryoET STA) has emerged as a potent tool to obtain structures of these at near-atomic resolution. The study of such complexes and assemblies in situ using cryoET STA, coupled with cryoFIB/SEM and correlative and integrative imaging, opens a new frontier in structural cell biology [2].  I will present our recent studies on human viruses, including HIV-1, SARS-CoV-2, and reovirus [3-5], to demonstrate the power of high-resolution cryoEM and in situ structural biology using cell lamellae-based cryoET STA.

References

[1] Henderson, R. (2009). Quarterly Reviews of Biophysics 28, 171.

[2] Zhang, P. (2019) Curr Opin Struct Biol 58, 249-258

[3] Sutton, G. (2020). Nat Commun 11(1):4445

[4] Mendonça, L. et al. (2021). Nat Commun 12(1):4629 

[5] Ni, T. et al. (2022). Sci Adv 7(47):eabj5715