Mutations in The Stator Protein PomA Affect Switching of Rotational Direction in Bacterial Flagellar Motor

Abstract The flagellar motor rotates bi-directionally in counter-clockwise (CCW) and clockwise (CW) directions. The motor consists of a stator and a rotor. Recent structural studies have revealed that the stator is composed of a pentameric ring of A subunits and a dimer axis of B subunits. The stator interacts with the rotor through conserved charged and neighboring residues, and the rotational power is generated by their interactions through a gear-like mechanism. The rotational direction is controlled by chemotaxis signaling transmitted to the rotor, with no evidence for the stator being involved. In this study, we found novel mutations that affect the switching of the rotational direction at the putative interaction site of the stator to generate rotational force. Our results highlight a novel aspect of flagellar motor function that appropriate switching of the interaction states between the stator and rotor is critical for controlling the rotational direction.

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Spontaneous adaptation of ion selectivity in a bacterial flagellar motor

10.1101/2021.01.26.427765 ◽

2021 ◽

Author(s):

Pietro Ridone ◽

Tsubasa Ishida ◽

Yoshiyuki Sowa ◽

Matthew A. B. Baker

Keyword(s):

Genome Engineering ◽

Environmental Changes ◽

Ion Selectivity ◽

Bacterial Species ◽

Selective Advantage ◽

Flagellar Motor ◽

Novel Mutations ◽

Power Sources ◽

Motor Complex ◽

Bacterial Flagellar Motor

ABSTRACTMotility provides a selective advantage to many bacterial species and is often achieved by rotation of flagella that propel the cell towards more favourable conditions. In most species, the rotation of the flagellum, driven by the Bacterial Flagellar Motor (BFM), is powered by H+ or Na+ ion transit through the torque-generating stator subunits of the motor complex. The ionic requirements for motility appear to have adapted to environmental changes throughout history but the molecular basis of this adaptation, and the constraints which govern the evolution of the stator proteins are unknown. Here we use CRISPR-mediated genome engineering to replace the native H+-powered stator genes of Escherichia coli with a compatible sodium-powered stator set from Vibrio alginolyticus and subsequently direct the evolution of the stators to revert to H+-powered motility. Evidence from whole genome sequencing indicates both flagellar- and non-flagellar-associated genes that are involved in longer-term adaptation to new power sources. Overall, transplanted Na+-powered stator genes can spontaneously incorporate novel mutations that allow H+-motility when environmental Na+ is lacking.

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Bacterial Flagellar Motor: Biochemical and Structural Studies

Encyclopedia of Biophysics ◽

10.1007/978-3-642-16712-6_198 ◽

2013 ◽

pp. 149-155

Author(s):

David Blair

Keyword(s):

Flagellar Motor ◽

Structural Studies ◽

Bacterial Flagellar Motor

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3P176 Pressure-induced reversal in the rotational direction of the bacterial flagellar motor(Molecular motors,Oral Presentations)

Seibutsu Butsuri ◽

10.2142/biophys.47.s247_1 ◽

2007 ◽

Vol 47 (supplement) ◽

pp. S247

Author(s):

Masayoshi Nishiyama ◽

Yoshiyuki Sowa ◽

Shigeichi Kumazaki ◽

Yoshifumi Kimura ◽

Michio Homma ◽

...

Keyword(s):

Molecular Motors ◽

Flagellar Motor ◽

Bacterial Flagellar Motor ◽

Oral Presentations ◽

Rotational Direction

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Molecular Mechanism for Rotational Switching of the Bacterial Flagellar Motor

10.1101/2020.05.18.101634 ◽

2020 ◽

Cited By ~ 1

Author(s):

Yunjie Chang ◽

Kai Zhang ◽

Brittany L. Carroll ◽

Xiaowei Zhao ◽

Nyles W. Charon ◽

...

Keyword(s):

Molecular Mechanism ◽

Conformational Changes ◽

Proton Motive Force ◽

Model System ◽

Dependent Manner ◽

Flagellar Motor ◽

Bacterial Flagellar Motor ◽

Rotational Direction ◽

Cytoplasmic Side

AbstractThe bacterial flagellar motor is a remarkable nanomachine that can rapidly rotate in both counter-clockwise (CCW) and clockwise (CW) senses. The transitions between CCW and CW rotation are critical for chemotaxis, and they are controlled by a signaling protein (CheY-P) that interacts with a switch complex at the cytoplasmic side of the flagellar motor. However, the exact molecular mechanism by which CheY-P controls the motor rotational switch remains enigmatic. Here, we use the Lyme disease spirochete, Borrelia burgdorferi, as the model system to dissect the mechanism underlying flagellar rotational switching. We first determined high resolution in situ motor structures in the cheX and cheY3 mutants in which motors are genetically locked in CCW or CW rotation. The structures showed that the CheY3 protein of B. burgdorferi interacts directly with the FliM protein of the switch complex in a phosphorylation-dependent manner. The binding of CheY3-P to FliM induces a major remodeling of the switch protein FliG2 that alters its interaction with the torque generator. Because the remodeling of FliG2 is directly correlated with the rotational direction, our data lead to a model for flagellar function in which the torque generator rotates in response to an inward flow of H+ driven by the proton motive force. Rapid conformational changes of FliG2 allow the switch complex to interact with opposite sides of the rotating torque generator, thereby facilitating rotational switching between CW and CCW.

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The flagellar motor of Vibrio alginolyticus undergoes major structural remodeling during rotational switching

eLife ◽

10.7554/elife.61446 ◽

2020 ◽

Vol 9 ◽

Cited By ~ 1

Author(s):

Brittany L Carroll ◽

Tatsuro Nishikino ◽

Wangbiao Guo ◽

Shiwei Zhu ◽

Seiji Kojima ◽

...

Keyword(s):

Conformational Changes ◽

Electron Tomography ◽

Protein Composition ◽

Vibrio Alginolyticus ◽

Flagellar Motor ◽

Conformational Rearrangement ◽

Structural Remodeling ◽

Cryo Electron Tomography ◽

Bacterial Flagellar Motor ◽

Rotational Direction

The bacterial flagellar motor switches rotational direction between counterclockwise (CCW) and clockwise (CW) to direct the migration of the cell. The cytoplasmic ring (C-ring) of the motor, which is composed of FliG, FliM, and FliN, is known for controlling the rotational sense of the flagellum. However, the mechanism underlying rotational switching remains elusive. Here, we deployed cryo-electron tomography to visualize the C-ring in two rotational biased mutants in Vibrio alginolyticus. We determined the C-ring molecular architectures, providing novel insights into the mechanism of rotational switching. We report that the C-ring maintained 34-fold symmetry in both rotational senses, and the protein composition remained constant. The two structures show FliG conformational changes elicit a large conformational rearrangement of the rotor complex that coincides with rotational switching of the flagellum. FliM and FliN form a stable spiral-shaped base of the C-ring, likely stabilizing the C-ring during the conformational remodeling.

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