In-cell NMR: a topical review.

Enrico Luchinat,Lucia Banci

doi:10.1107/s2052252516020625

Enrico Luchinat, Lucia Banci

Open Access

https://doi.org/10.1107/s2052252516020625

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Abstract

Classical structural biology approaches allow structural characterization of biological macromolecules in vitro, far from their physiological context. Nowadays, thanks to the wealth of structural data available and to technological and methodological advances, the interest of the research community is gradually shifting from pure structural determination towards the study of functional aspects of biomolecules. Therefore, a cellular structural approach is ideally needed to characterize biological molecules, such as proteins, in their native cellular environment and the functional processes that they are involved in. In-cell NMR is a new application of high-resolution nuclear magnetic resonance spectroscopy that allows structural and dynamical features of proteins and other macromolecules to be analyzed directly in living cells. Owing to its challenging nature, this methodology has shown slow, but steady, development over the past 15 years. To date, several in-cell NMR approaches have been successfully applied to both bacterial and eukaryotic cells, including several human cell lines, and important structural and functional aspects have been elucidated. In this topical review, the major advances of in-cell NMR are summarized, with a special focus on recent developments in eukaryotic and mammalian cells.

Highlights

The structure of biological macromolecules is critical to understanding their function, their mode of interaction and relation with their partners, and how physiological processes are altered by mutations or changes in the molecular environment
We summarize the major advances of in-cell nuclear magnetic resonance (NMR) since its first application, and we further report the recent developments of this promising methodology, with a special focus on its application to study proteins in eukaryotic and mammalian cells and on the development of cellular solid-state NMR approaches
Serber and coworkers showed that small globular proteins could be overexpressed in E. coli and isotopically labelled to a sufficient level that it was possible to detect them above the other cellular components by heteronuclear NMR (Serber et al, 2001)

Summary

Introduction

The structure of biological macromolecules is critical to understanding their function, their mode of interaction and relation with their partners, and how physiological processes are altered by mutations or changes in the molecular environment. Being especially suited to investigate the structure and dynamics of macromolecules at atomic resolution, in-cell NMR can fill a critical gap between in vitro-oriented structural techniques, such as NMR spectroscopy, X-ray crystallography and single-particle cryo-EM, and ultrahighresolution cellular imaging techniques, such as superresolution microscopy and cryo-electron tomography, which have seen impressive development in recent years. In this topical review, we summarize the major advances of in-cell NMR since its first application, and we further report the recent developments of this promising methodology, with. A special focus on its application to study proteins in eukaryotic and mammalian cells and on the development of cellular solid-state NMR approaches

Overview of in-cell NMR approaches

Cellular solid-state NMR and DNP

Solution structure determination in living cells

Biological insights

Findings

Future perspectives