CryoEM structures reveal how the bacterial flagellum rotates and switches direction

Prashant K. Singh; Pankaj Sharma; Oshri Afanzar; Margo H. Goldfarb; Elena Maklashina; Michael Eisenbach; Gary Cecchini; T. M. Iverson

doi:10.1038/s41564-024-01674-1

CryoEM structures reveal how the bacterial flagellum rotates and switches direction

Prashant K. Singh, Pankaj Sharma, Oshri Afanzar, Margo H. Goldfarb, Elena Maklashina, Michael Eisenbach, Gary Cecchini, T. M. Iverson^*

^*Corresponding author for this work

Department of Biomolecular Sciences

Research output: Contribution to journal › Article › peer-review

3 Citations (Scopus)

Abstract

Bacterial chemotaxis requires bidirectional flagellar rotation at different rates. Rotation is driven by a flagellar motor, which is a supercomplex containing multiple rings. Architectural uncertainty regarding the cytoplasmic C-ring, or ‘switch’, limits our understanding of how the motor transmits torque and direction to the flagellar rod. Here we report cryogenic electron microscopy structures for Salmonella enterica serovar typhimurium inner membrane MS-ring and C-ring in a counterclockwise pose (4.0 Å) and isolated C-ring in a clockwise pose alone (4.6 Å) and bound to a regulator (5.9 Å). Conformational differences between rotational poses include a 180° shift in FliF/FliG domains that rotates the outward-facing MotA/B binding site to inward facing. The regulator has specificity for the clockwise pose by bridging elements unique to this conformation. We used these structures to propose how the switch reverses rotation and transmits torque to the flagellum, which advances the understanding of bacterial chemotaxis and bidirectional motor rotation.

Original language	English
Pages (from-to)	1271-1281
Number of pages	11
Journal	Nature Microbiology
Volume	9
Issue number	5
DOIs	https://doi.org/10.1038/s41564-024-01674-1
Publication status	Published Online - 17 Apr 2024

Bibliographical note

We thank S. Khan, currently at the Molecular Biology Consortium, Lawrence Berkley National Laboratories and Lahore University of Management Sciences, for the pKLR3 plasmid; T. Nakagawa for technical advice during the early stages of this work; and B. Butler for experimental assistance. We thank W. Chiu and the S2C2 workshop for training in cryoEM. We thank B. DeBuyser for the artificial intelligence-based goblin.tools. This project was funded by National Institutes of Health (NIH) grant GM61606 awarded to T.M.I. and G.C. G.C. is the recipient of a Senior Research Career Scientist award 1K6BX004215 from the Department of Veterans Affairs. M.H.G. is supported by NIH T32 GM007628. Negative stain and cryoEM data were collected at the Center for Structural Biology CryoEM facility at Vanderbilt University. The Glacios cryo-TEM used for screening was acquired by NIH grant S10 OD030292-01.

Publisher Copyright:
© The Author(s) 2024.

All Science Journal Classification (ASJC) codes

Microbiology
Immunology
Applied Microbiology and Biotechnology
Genetics
Microbiology (medical)
Cell Biology

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Cite this

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title = "CryoEM structures reveal how the bacterial flagellum rotates and switches direction",

abstract = "Bacterial chemotaxis requires bidirectional flagellar rotation at different rates. Rotation is driven by a flagellar motor, which is a supercomplex containing multiple rings. Architectural uncertainty regarding the cytoplasmic C-ring, or {\textquoteleft}switch{\textquoteright}, limits our understanding of how the motor transmits torque and direction to the flagellar rod. Here we report cryogenic electron microscopy structures for Salmonella enterica serovar typhimurium inner membrane MS-ring and C-ring in a counterclockwise pose (4.0 {\AA}) and isolated C-ring in a clockwise pose alone (4.6 {\AA}) and bound to a regulator (5.9 {\AA}). Conformational differences between rotational poses include a 180° shift in FliF/FliG domains that rotates the outward-facing MotA/B binding site to inward facing. The regulator has specificity for the clockwise pose by bridging elements unique to this conformation. We used these structures to propose how the switch reverses rotation and transmits torque to the flagellum, which advances the understanding of bacterial chemotaxis and bidirectional motor rotation.",

author = "Singh, {Prashant K.} and Pankaj Sharma and Oshri Afanzar and Goldfarb, {Margo H.} and Elena Maklashina and Michael Eisenbach and Gary Cecchini and Iverson, {T. M.}",

note = "We thank S. Khan, currently at the Molecular Biology Consortium, Lawrence Berkley National Laboratories and Lahore University of Management Sciences, for the pKLR3 plasmid; T. Nakagawa for technical advice during the early stages of this work; and B. Butler for experimental assistance. We thank W. Chiu and the S2C2 workshop for training in cryoEM. We thank B. DeBuyser for the artificial intelligence-based goblin.tools. This project was funded by National Institutes of Health (NIH) grant GM61606 awarded to T.M.I. and G.C. G.C. is the recipient of a Senior Research Career Scientist award 1K6BX004215 from the Department of Veterans Affairs. M.H.G. is supported by NIH T32 GM007628. Negative stain and cryoEM data were collected at the Center for Structural Biology CryoEM facility at Vanderbilt University. The Glacios cryo-TEM used for screening was acquired by NIH grant S10 OD030292-01. Publisher Copyright: {\textcopyright} The Author(s) 2024.",

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AU - Cecchini, Gary

AU - Iverson, T. M.

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N2 - Bacterial chemotaxis requires bidirectional flagellar rotation at different rates. Rotation is driven by a flagellar motor, which is a supercomplex containing multiple rings. Architectural uncertainty regarding the cytoplasmic C-ring, or ‘switch’, limits our understanding of how the motor transmits torque and direction to the flagellar rod. Here we report cryogenic electron microscopy structures for Salmonella enterica serovar typhimurium inner membrane MS-ring and C-ring in a counterclockwise pose (4.0 Å) and isolated C-ring in a clockwise pose alone (4.6 Å) and bound to a regulator (5.9 Å). Conformational differences between rotational poses include a 180° shift in FliF/FliG domains that rotates the outward-facing MotA/B binding site to inward facing. The regulator has specificity for the clockwise pose by bridging elements unique to this conformation. We used these structures to propose how the switch reverses rotation and transmits torque to the flagellum, which advances the understanding of bacterial chemotaxis and bidirectional motor rotation.

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CryoEM structures reveal how the bacterial flagellum rotates and switches direction

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