The advent of structural biology in situ by single particle cryo-electron tomography

Jesús G Galaz-Montoya,Steven J Ludtke

doi:10.1007/s41048-017-0040-0

Jesús G Galaz-Montoya, Steven J Ludtke

Open Access

https://doi.org/10.1007/s41048-017-0040-0

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Journal: Biophysics reports	Publication Date: May 29, 2017
Citations: 43	License type: open-access

Affiliation: Baylor College of Medicine

Abstract

Single particle tomography (SPT), also known as subtomogram averaging, is a powerful technique uniquely poised to address questions in structural biology that are not amenable to more traditional approaches like X-ray crystallography, nuclear magnetic resonance, and conventional cryoEM single particle analysis. Owing to its potential for in situ structural biology at subnanometer resolution, SPT has been gaining enormous momentum in the last five years and is becoming a prominent, widely used technique. This method can be applied to unambiguously determine the structures of macromolecular complexes that exhibit compositional and conformational heterogeneity, both in vitro and in situ. Here we review the development of SPT, highlighting its applications and identifying areas of ongoing development.

Highlights

THE NEED FOR SINGLE PARTICLE TOMOGRAPHYOver the last five years, thanks to the development of direct detection devices (DDDs) (Milazzo et al 2005), cryo-electron microscopy single particle analysis (SPA) has transitioned from being an established, but limited, technique to being at the forefront of structural biology (Eisenstein 2016; Nogales 2016)
The iterative compressed-sensing optimized non-uniform (ICON) reconstruction (Deng et al 2016) method demonstrated that CS can restore missing information in noisy cryo-electron tomography (cryoET) data of both cells and isolated macromolecules, thereby minimizing missing wedge artifacts and yielding measurably better reconstructions than weighted back-projection (WBP)
Some algorithms rely on the individual images in a tiltseries having high contrast, such as the filtered iterative reconstruction technique (FIRT) (Chen et al 2016), which does not seem to provide any advantages over WBP when applied to cryoET data

Summary

Introduction

THE NEED FOR SINGLE PARTICLE TOMOGRAPHYOver the last five years, thanks to the development of direct detection devices (DDDs) (Milazzo et al 2005), cryo-electron microscopy (cryoEM) single particle analysis (SPA) has transitioned from being an established, but limited, technique to being at the forefront of structural biology (Eisenstein 2016; Nogales 2016). The recent development of direct electron detectors, phase plate technology, and improved contrast transfer function (CTF) correction methodologies for cryoET have made it possible to achieve images with higher contrast and resolution, respectively, using much lower dose.

Results

Conclusion