Complete structure of the chemosensory array core signalling unit in an E. coli minicell strain

Alister Burt,Maria Bacia-Verloop,John S Parkinson,Irina Gutsche,Ambroise Desfosses,Megghane Baulard,Karine Huard,Phillip J Stansfeld,C Keith Cassidy,William Margolin,Peter Ames,Zaida Luthey-Schulten

doi:10.1038/s41467-020-14350-9

Abstract

Motile bacteria sense chemical gradients with transmembrane receptors organised in supramolecular signalling arrays. Understanding stimulus detection and transmission at the molecular level requires precise structural characterisation of the array building block known as a core signalling unit. Here we introduce an Escherichia coli strain that forms small minicells possessing extended and highly ordered chemosensory arrays. We use cryo-electron tomography and subtomogram averaging to provide a three-dimensional map of a complete core signalling unit, with visible densities corresponding to the HAMP and periplasmic domains. This map, combined with previously determined high resolution structures and molecular dynamics simulations, yields a molecular model of the transmembrane core signalling unit and enables spatial localisation of its individual domains. Our work thus offers a solid structural basis for the interpretation of a wide range of existing data and the design of further experiments to elucidate signalling mechanisms within the core signalling unit and larger array.

Highlights

Motile bacteria sense chemical gradients with transmembrane receptors organised in supramolecular signalling arrays
Cryo-electron tomography structures of complexes composed of receptor cytoplasmic domains, CheA and CheW that are assembled on lipid monolayers provide insights into core-signalling units (CSUs) structure[9], but lack the ligandbinding, transmembrane, and HAMP portions of the receptor
While the primary aim of this work was to solve the complete structure of a single CSU, our approach may be extended to the higher-order array structure

Summary

Introduction

Motile bacteria sense chemical gradients with transmembrane receptors organised in supramolecular signalling arrays. Changes in the occupancy of MCP periplasmic ligand-binding domains trigger conformational rearrangements that propagate through the inner membrane to the HAMP domain (a signalling element which couples extracellular input to intracellular output in most microbial chemoreceptors and sensory kinases)[2] and through an extended four-helix cytoplasmic domain[1] The latter comprises three signaling regions: (1) a methylation-helix (MH) bundle, which contains the sites of reversible modification, (2) a flexible region containing the glycine hinge[3], and (3) the kinasecontrol region that binds CheA and CheW and enables receptor trimerisation (Fig. 1a). We present the WM4196 minicell-producing E. coli strain from which we isolated small, healthy-looking minicells suitable for high-resolution analysis of internal structures by cryo-ET and subtomogram averaging We observe that these cells contain extended, ordered and functional chemosensory arrays and can be used to revisit the structure of the CSU. The resulting three-dimensional (3D) map enables us to propose a molecular model of the transmembrane CSU, which opens up avenues for further research

Objectives

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Conclusion