Bacteria are so small that for them, making their way through water is like swimming in roofing tar for us. In response, these organisms have evolved a molecular machine that helps them move in their environment. Named the bacterial flagellum, this complex assemblage of molecules is formed of three main parts: a motor that spans the inner and outer membranes of the cell, and then a ‘hook’ that connects to a long filament which extends outside the bacterium. More precisely, the motor is formed of the stator, an ion pump that stays still, and of a rotor that can spin. Different rings can also be present in the space between the inner and outer membranes (the periplasm) and surround these components. The stator uses ions to generate the energy that makes the rotor whirl. In turn, this movement sets the filament in motion, propelling the bacterium. Depending on where the bacteria live, the stator can use different types of ions. In addition, while many species have a single stator system per motor, some may have several stator systems for one motor: this may help the microorganisms move in different conditions.

As microbes colonize environments with a different pH or viscosity, they constantly evolve new versions of the motor which are more suitable to their new surroundings. However, a part of the motor remains the same across species. Overall, it is still unclear how bacterial flagella evolve, but examining the structure of new motors can shed light upon this process. Here, Kaplan et al. combine a bioinformatics approach with an imaging technique known as electron cryo-tomography to dissect the structure of the flagellar motor of three species of bacteria with different stator systems, and compare these to known motors of the same class.

The results reveal a correlation between the nature of the stator system and the presence of certain elements. Stators that use sodium ions, or both sodium and hydrogen ions, are associated with two periplasmic rings surrounding the conserved motor structure. These rings do not exist in motors with single hydrogen-driven stators. Motors with dual hydrogen-driven stators are, to some extent, an ‘intermediate state’, with only one of those rings present. As all the studied species currently exist, it is difficult to know which version of the motor is the most ancient, and which one has evolved more recently.

Capturing the diversity of bacterial motors gives us insight into the evolutionary forces that shape complex molecular structures, which is essential to understand how life evolved on Earth. More practically, this knowledge may also help us design better nanomachines to power microscopic robots.

The bacterial flagellum is a macromolecular machine that transforms the movement of ions (H⁺, Na⁺ or other cations) across the cell membrane into a mechanical torque to move the bacterial cell through its environment (Ito and Takahashi, 2017; Sowa and Berry, 2008). In general, the flagellum consists of a cell-envelope-embedded motor, a hook which acts as a universal joint and a long propeller-like filament (Berg, 2003; Erhardt et al., 2010). The motor is composed of a rotor and a stator: while the stator is the part of the motor that remains static, the rotor is the part that rotates and can rotate the filament in either a counterclockwise or clockwise direction. For cells with a single flagellum this drives the cell forward or backward; for peritrichous cells this results in ‘run’ or ‘tumble’ movements. Flagella can also exhibit a more complex behavior; it was recently reported that the Shewanella putrefaciens flagellum can wrap around the cell to mediate a screw-like motion that allows the cell to escape narrow traps (Kühn et al., 2017). Besides their role in motility, bacterial flagella participate in other vital activities of the cell such as biofilm formation (Belas, 2014). Moreover, the virulence of many human pathogens depends directly on their flagella, with flagellated strains of Pseudomonas aeruginosa and Legionella pneumophila causing more serious infections with higher mortality rates (Appelt and Heuner, 2017; Feldman et al., 1998). P. aeruginosa lacking fully-assembled flagella cause no mortality and are 75% less likely to cause pneumonia in mice (Feldman et al., 1998).

The best-studied flagellar motor, in Salmonella enterica, consists of several sub-complexes, which we will describe in order from the inside out. On the cytoplasmic side are the inner-membrane-embedded MS ring (formed by the protein FliF) and the C-ring (aka the switch complex, formed by FliN, FliM and FliG). The C-ring encircles a type III secretion system (T3SS) export apparatus (FliH, FliI, FliJ, FlhA, FlhB, FliP, FliQ and FliR). Spanning the space from the inner membrane to the peptidoglycan cell wall is the ion channel (called the stator), a complex of two proteins (MotA and MotB) with 4:2 stoichiometry (Koebnik, 1995; Kojima, 2015; Morimoto and Minamino, 2014). This complex is anchored to the peptidoglycan and converts the flux of ions across the bacterial membrane into a torque through the interaction of the so-called ‘torque-helix’ in MotA with FliG in the C-ring. Previous studies have shown that cycles of protonation/deprotonation of a certain aspartate residue in the cytoplasmic end of MotB induce conformational changes in MotA, which in turn interacts with the C-terminus of FliG. How the torque that is generated through this interaction is transferred to the other parts of the motor remains unclear with different models suggested (see Berg, 2003; Stock et al., 2012 and references therein for details). The MS ring is coupled to the extracellular hook (FlgE) through the rod (FlgB, FlgC, FlgF and FlgG). The rod is further surrounded by two other rings: the P- (peptidoglycan, FlgI) and the L- (lipopolysaccharide, FlgH) rings which act as bushings during rod rotation. Extending from the hook is the filament (FliC) which is many micrometers in length. In addition to these components, the assembly of the whole flagellar motor is a highly synchronized process that requires a plethora of additional chaperones and capping proteins (Altegoer and Bange, 2015; Evans et al., 2014; Jones and Macnab, 1990; Kaplan et al., 2018; Kubori et al., 1992; Macnab, 1999).

Recently, the development of electron cryo-tomography (ECT) (Gan and Jensen, 2012; Oikonomou and Jensen, 2017; Pfeffer and Förster, 2018) has allowed the determination of the complete structures of flagellar motors in their cellular milieu at macromolecular (~5 nm) resolution. ECT studies of many different bacterial species have revealed that while the core structure described above is conserved, the flagellar motor has evolved many species-specific adaptations to different environmental conditions (Beeby et al., 2016; Chaban et al., 2018; Chen et al., 2011; Minamino and Imada, 2015; Terashima et al., 2017; Zhao et al., 2014; Zhu et al., 2017). For example, extra periplasmic rings were found to elaborate the canonical P- and L-rings in the motor of the Gammaproteobacteria Vibrio species. These rings are called the T-ring (MotX and Y) and H-ring (FlgO, P and T) (Terashima et al., 2010; Terashima et al., 2006). Unlike the S. enterica motor described above, which is driven by H⁺ ions, the motors of Vibrio and other marine bacteria employ different stators (PomA and PomB) which utilize Na⁺. These Na⁺-dependent stators generate higher torque (~2200 pN) than H⁺-dependent stators (~1200 pN), driving the motor at higher speeds (up to 1,700 Hz compared to ~300 Hz in H⁺-driven motors) (Magariyama et al., 1994).

Many flagellated bacteria use a single stator system – either H⁺-driven or Na⁺-driven, depending on their environment. Interestingly, it has also been shown that the single stator system of Bacillus clausii KSM-K16 is able to use both Na⁺ and H⁺ at different pH levels (Terahara et al., 2008). Additionally, some species (like alkaliphilic Bacillus alcalophilus AV1934 and Paenibacillus sp. TCA20), can use other cations to generate the energy required for torque generation depending on their environment (Ito and Takahashi, 2017; Terahara et al., 2012). Some species, however, such as Vibrio alginolyticus, use two distinct types of motors to move in different environments: a polar Na⁺-driven flagellum and lateral H⁺-driven flagella. Still other species employ dual stator systems with a single flagellar motor, conferring an advantage for bacteria that experience a range of environments (see Thormann and Paulick, 2010 and references therein). For example, P. aeruginosa employs a dual H⁺-driven stator system (MotAB and MotCD). While the MotAB system is sufficient to move the cell in a liquid environment (Doyle et al., 2004), MotCD is necessary to allow the cell to move in more viscous conditions (Toutain et al., 2007). Shewanella oneidensis MR-1 combines both Na⁺- and H⁺-dependent stators in a single motor, enabling the bacterium to move efficiently under conditions of different pH and Na⁺ concentration (Paulick et al., 2015). How these more elaborate motors may have evolved remains an open question.

Here, we used ECT to determine the first in situ structures of three Gammaproteobacteria flagellar motors with dual stator systems: in L. pneumophila, P. aeruginosa and S. oneidensis MR-1. L. pneumophila and P. aeruginosa have dual H⁺-dependent stator systems and S. oneidensis has a dual Na⁺-H⁺-dependent stator. This imaging, along with bioinformatics analysis, shows a correlation between the structural elaboration of the motor and its stator system, suggesting a possible evolutionary pathway.

To determine the structures of the flagellar motors of L. pneumophila, P. aeruginosa, and S. oneidensis we imaged intact cells of each species in a hydrated frozen state using ECT. We identified clearly visible flagellar motors in the tomographic reconstructions and performed sub-tomogram averaging to enhance the signal-to-noise ratio, generating a 3D average of the motor of each species at macromolecular resolution (Figure 1 and Figure 1—figure supplement 1). Although the three motors shared the conserved core structure of the flagellar motor, they exhibited different periplasmic decorations surrounding this conserved core. While the S. oneidensis and P. aeruginosa averages showed clear densities corresponding to the stators (Figure 1E,F,K and L, dark orange density), none were visible in the L. pneumophila average, suggesting that they were more variable, or dynamic and therefore are not visible in the average (see e.g., (Chen et al., 2011; Zhu et al., 2017)). Interestingly, we observed a novel feature in the S. oneidensis motor: an extra ring outside the outer membrane (Figure 1A–F, purple density). Although in some tomograms two extracellular rings appeared to be present (see Figure 1A and C), only one ring was visible in the sub-tomogram average which could be either because one of the rings is more dynamic or substoichiometric (Figure 1B,D and Figure 1—figure supplement 2 for more examples of single motors). This structure is reminiscent of the O-ring (outer membrane ring) described recently in the sheathed flagellum of Vibrio alginolyticus (Zhu et al., 2017). However, while the V. alginolyticus O-ring was associated with a 90° bend in the outer membrane, no such outer membrane bend was seen in the unsheathed S. oneidensis flagellum, so the function of this structure remains mysterious.

Figure 1 with 2 supplements see all

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The most striking difference between the three motor structures was the L- and P-rings, which were highly elaborated in S. oneidensis. The P. aeruginosa and L. pneumophila motors lacked additional rings associated with the L-ring, but showed smaller elaborations of their P-rings. To determine whether flagellar motor structure correlates with stator type, we compared our three new ECT structures with those of the five previously-published Gammaproteobacteria motors (Figure 2). Two motors (Escherichia coli and S. enterica) have a single H⁺-driven stator system, two motors have dual H⁺-dependent stator systems (P. aeruginosa and L. pneumophila), three motors have Na⁺-driven systems (the three Vibrio species) and one motor has a dual Na⁺-H⁺-driven system (S. oneidensis). Interestingly, we found that motors with similar stator type also shared similar structural characteristics. While the two motors with a single H⁺-dependent stator system did not show any periplasmic elaborations beyond the conserved flagellar core, the dual H⁺-dependent stator systems had an extra ring surrounding their P-ring, with no embellishment of the L-ring. The Na⁺-dependent motors of the Vibrio spp., together with the Na⁺-H⁺-dependent motor of S. oneidensis, have extra components surrounding both their P- and L- rings. In Vibrio, these extra periplasmic rings are known as the T-ring (surrounding the P- ring and formed by the MotX and MotY proteins) and the H-ring (surrounding the L-ring and consisting of the FlgO, FlgP and FlgT proteins). The presence of the T- and H-rings was suggested to be specific to the Na⁺-driven Vibrio motors (Minamino and Imada, 2015) with the FlgT protein required for the formation of both rings (Terashima et al., 2013).

Figure 2

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Previous studies showed that MotX and MotY are important for flagellar rotation in S. oneidensis but it was not known whether they form part of the motor or not (Koerdt et al., 2009). Similarly, bioinformatics analysis and biochemical studies showed that MotY is involved in the function of the P. aeruginosa motor, but the structural basis of this role was not known (Doyle et al., 2004). We therefore performed a bioinformatics search for candidate homologs of MotX, MotY, FlgO, FlgP and FlgT in the genomes of P. aeruginosa, L. pneumophila and S. oneidensis to examine whether there is a correlation between the presence of homologous genes and the extra periplasmic rings observed in the ECT structures. While we found candidates for all five proteins constituting the T- and H-rings in S. oneidensis as previously suggested (Wu et al., 2011), only MotY candidates were found in L. pneumophila and P. aeruginosa (Table 1). This is in accordance with our ECT structures, which showed that L. pneumophila and P. aeruginosa motors have a ring surrounding only their P-rings while the S. oneidensis motor has rings surrounding both the P- and L-rings. These rings are likely T- and H-rings, respectively, as in Vibrio. The lack of candidate MotX homologs in the genomes of L. pneumophila and P. aeruginosa (Table 1) is consistent with their lack of PomB, the component of the Na⁺-dependent stator with which MotX interacts. Interestingly, the absence of candidates for FlgT in the L. pneumophila and P. aeruginosa genomes suggests that it may not be required for the recruitment of MotY as in Vibrio species.

To see whether these correlations hold more broadly, we expanded our bioinformatics analysis to additional species of Gammaproteobacteria (Williams et al., 2010). We examined the genomes of species with single H⁺-driven stator systems (Table 2), dual H⁺-driven stator systems (Table 3) and Na⁺-driven stator systems (Table 4). These species were identified either by Blasting the sequence of the stator proteins (MotA, B, C and D and Pom A and B, see Materials and methods) against the genome of the species or based on previous studies (Thormann and Paulick, 2010). In all species we examined, we observed the same pattern: (i) genomes of species with single H⁺-driven stator systems lacked homologs of H- or T-ring components; (ii) genomes of species with Na⁺ (or Na⁺-H⁺) stator systems contained homologs of all H- and T-ring components, and (iii) genomes of species with dual H⁺-driven stator systems contained candidate homologs only for the T-ring component MotY. The sole exception to this rule was Chromohalobacter salexigens DSM 3043, which contained a homolog of FlgO in addition to MotY. Also, while Serratia proteomaculans and Psychromonas ingrahamii have candidates for single MotAB stator system they also have candidates for MotY (see Supplementary file 1 and 2). None of the thirteen species with dual H⁺-driven stator systems we examined contained a homolog of FlgT, further suggesting that it is not essential for MotY stabilization in this group.

Together, our results from ECT imaging of flagellar motors in situ and bioinformatics analysis reveal a correlation between the structural elaboration of the flagellar motor of Gammaproteobacteria and the type of its torque-generating unit, the stator (summarized in Figure 3). Low-speed motors with single H⁺-stator systems have only the P- and L-rings, while high-speed motors using Na⁺ have two extra periplasmic rings, the T- and H-rings. Unexpectedly, we find that motors with dual H⁺-driven stator systems represent a hybrid structure between the two, elaborating their P-rings with one of the five components of the T- and H-rings, MotY. It is important to note that the presence of these extra elaborations in the motor is encoded in the genome and is not related to whether or not a stator subunit is recruited on the motor. This extra MotY ring might help to stabilize the motor under conditions of increased load, as in the viscous environment of the pulmonary system encountered by L. pneumophila and P. aeruginosa. These results therefore suggest an evolutionary pathway in which these pathogenic Gammaproteobacteria species could have borrowed a motor stabilization strategy from related Na⁺-driven motors to allow them to colonize animal hosts. Finally, It would be interesting to investigate whether our observation here holds for other bacterial species that use different cations as their energy source (Ito and Takahashi, 2017) and whether it extends to other bacterial species with more than two stator systems or other classes.

Figure 3

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All data generated or analysed during this study are included in the manuscript and supporting files. The ECT structures have been deposited in the EMDB under the following accession numbers, EMD-0464 for Legionella pneumophila motor, EMD-0465 for Pseudomonas aeruginosa motor and EMD-0467 for Shewanella oneidensis MR-1 motor.

1. 1. Mohammed Kaplan
   2. Debnath Ghosal
   3. Poorna Subramanian
   4. Catherine M Oikonomou
   5. Andreas Kjaer
   6. Sahand Pirbadian
   7. Davi R Ortega
   8. Ariane Briegel
   9. Mohamed Y El-Naggar
   10. Grant J Jensen

   (2019) EM Data Bank

   ID EMD-0464. Legionella pneumophila motor.

   http://www.ebi.ac.uk/pdbe/entry/emdb/EMD-0464
2. 1. Mohammed Kaplan
   2. Debnath Ghosal
   3. Poorna Subramanian
   4. Catherine M Oikonomou
   5. Andreas Kjaer
   6. Sahand Pirbadian
   7. Davi R Ortega
   8. Ariane Briegel
   9. Mohamed Y El-Naggar
   10. Grant J Jensen

   (2019) EM Data Bank

   ID EMD-0467. Shewanella oneidensis MR-1 motor.

   http://www.ebi.ac.uk/pdbe/entry/emdb/EMD-0467
3. 1. Mohammed Kaplan
   2. Debnath Ghosal
   3. Poorna Subramanian
   4. Catherine M Oikonomou
   5. Andreas Kjaer
   6. Sahand Pirbadian
   7. Davi R Ortega
   8. Ariane Briegel
   9. Mohamed Y El-Naggar
   10. Grant J Jensen

   (2019) EM Data Bank

   ID EMD-0465. Pseudomonas aeruginosa motor.

   http://www.ebi.ac.uk/pdbe/entry/emdb/EMD-0465

1. 1. Agulleiro JI
   2. Fernandez JJ

   (2011) Fast tomographic reconstruction on multicore computers

   Bioinformatics 27:582–583.

   https://doi.org/10.1093/bioinformatics/btq692

   • PubMed
   • Google Scholar
2. 1. Altegoer F
   2. Bange G

   (2015) Undiscovered regions on the molecular landscape of flagellar assembly

   Current Opinion in Microbiology 28:98–105.

   https://doi.org/10.1016/j.mib.2015.08.011

   • PubMed
   • Google Scholar
3. 1. Appelt S
   2. Heuner K

   (2017) The flagellar regulon of Legionella-A Review

   Frontiers in Cellular and Infection Microbiology 7:454.

   https://doi.org/10.3389/fcimb.2017.00454

   • PubMed
   • Google Scholar
4. 1. Beeby M
   2. Ribardo DA
   3. Brennan CA
   4. Ruby EG
   5. Jensen GJ
   6. Hendrixson DR

   (2016) Diverse high-torque bacterial flagellar motors assemble wider stator rings using a conserved protein scaffold

   PNAS 113:E1917–E1926.

   https://doi.org/10.1073/pnas.1518952113

   • PubMed
   • Google Scholar
5. 1. Belas R

   (2014) Biofilms, flagella, and mechanosensing of surfaces by bacteria

   Trends in Microbiology 22:517–527.

   https://doi.org/10.1016/j.tim.2014.05.002

   • PubMed
   • Google Scholar
6. 1. Berg HC

   (2003) The rotary motor of bacterial flagella

   Annual Review of Biochemistry 72:19–54.

   https://doi.org/10.1146/annurev.biochem.72.121801.161737

   • PubMed
   • Google Scholar
7. 1. Bouhenni RA
   2. Vora GJ
   3. Biffinger JC
   4. Shirodkar S
   5. Brockman K
   6. Ray R
   7. Wu P
   8. Johnson BJ
   9. Biddle EM
   10. Marshall MJ
   11. Fitzgerald LA
   12. Little BJ
   13. Fredrickson JK
   14. Beliaev AS
   15. Ringeisen BR
   16. Saffarini DA

   (2010) The role of shewanella oneidensis MR-1 outer surface structures in extracellular electron transfer

   Electroanalysis 22:856–864.

   https://doi.org/10.1002/elan.200880006

   • Google Scholar
8. 1. Chaban B
   2. Coleman I
   3. Beeby M

   (2018) Evolution of higher torque in Campylobacter-type bacterial flagellar motors

   Scientific Reports 8:97.

   https://doi.org/10.1038/s41598-017-18115-1

   • PubMed
   • Google Scholar
9. 1. Charania MA
   2. Brockman KL
   3. Zhang Y
   4. Banerjee A
   5. Pinchuk GE
   6. Fredrickson JK
   7. Beliaev AS
   8. Saffarini DA

   (2009) Involvement of a membrane-bound class III adenylate cyclase in regulation of anaerobic respiration in shewanella oneidensis MR-1

   Journal of Bacteriology 191:4298–4306.

   https://doi.org/10.1128/JB.01829-08

   • PubMed
   • Google Scholar
10. 1. Chen S
    2. Beeby M
    3. Murphy GE
    4. Leadbetter JR
    5. Hendrixson DR
    6. Briegel A
    7. Li Z
    8. Shi J
    9. Tocheva EI
    10. Müller A
    11. Dobro MJ
    12. Jensen GJ

    (2011) Structural diversity of bacterial flagellar motors

    The EMBO Journal 30:2972–2981.

    https://doi.org/10.1038/emboj.2011.186

    • PubMed
    • Google Scholar
11. 1. Doyle TB
    2. Hawkins AC
    3. McCarter LL

    (2004) The complex flagellar torque generator of pseudomonas aeruginosa

    Journal of Bacteriology 186:6341–6350.

    https://doi.org/10.1128/JB.186.19.6341-6350.2004

    • PubMed
    • Google Scholar
12. 1. Erhardt M
    2. Namba K
    3. Hughes KT

    (2010) Bacterial nanomachines: the flagellum and type III injectisome

    Cold Spring Harbor Perspectives in Biology 2:a000299.

    https://doi.org/10.1101/cshperspect.a000299

    • PubMed
    • Google Scholar
13. 1. Evans LD
    2. Hughes C
    3. Fraser GM

    (2014) Building a flagellum outside the bacterial cell

    Trends in Microbiology 22:566–572.

    https://doi.org/10.1016/j.tim.2014.05.009

    • PubMed
    • Google Scholar
14. 1. Feldman M
    2. Bryan R
    3. Rajan S
    4. Scheffler L
    5. Brunnert S
    6. Tang H
    7. Prince A

    (1998)

    Role of flagella in pathogenesis of pseudomonas aeruginosa pulmonary infection

    Infection and Immunity 66:43–51.

    • PubMed
    • Google Scholar
15. 1. Gan L
    2. Jensen GJ

    (2012) Electron tomography of cells

    Quarterly Reviews of Biophysics 45:27–56.

    https://doi.org/10.1017/S0033583511000102

    • PubMed
    • Google Scholar
16. 1. Ito M
    2. Takahashi Y

    (2017) Nonconventional cation-coupled flagellar motors derived from the alkaliphilic Bacillus and paenibacillus species

    Extremophiles 21:3–14.

    https://doi.org/10.1007/s00792-016-0886-y

    • PubMed
    • Google Scholar
17. 1. Jones CJ
    2. Macnab RM

    (1990) Flagellar assembly in salmonella typhimurium: analysis with temperature-sensitive mutants

    Journal of Bacteriology 172:1327–1339.

    https://doi.org/10.1128/jb.172.3.1327-1339.1990

    • PubMed
    • Google Scholar
18. Preprint

    1. Kaplan M
    2. Subramanian P
    3. Ghosal D
    4. Oikonomou CM
    5. Pirbadian S
    6. Starwalt-Lee R
    7. Gralnick JA
    8. El-Naggar MY
    9. Jensen GJ

    (2018) Stable sub-complexes observed in situ suggest a modular assembly pathway of the bacterial flagellar motor

    bioRxiv.

    https://doi.org/10.1101/369405

    • Google Scholar
19. 1. Koebnik R

    (1995) Proposal for a peptidoglycan-associating alpha-helical motif in the C-terminal regions of some bacterial cell-surface proteins

    Molecular Microbiology 16:1269–1270.

    https://doi.org/10.1111/j.1365-2958.1995.tb02348.x

    • Google Scholar
20. 1. Koerdt A
    2. Paulick A
    3. Mock M
    4. Jost K
    5. Thormann KM

    (2009) MotX and MotY are required for flagellar rotation in Shewanella oneidensis MR-1

    Journal of Bacteriology 191:5085–5093.

    https://doi.org/10.1128/JB.00206-09

    • PubMed
    • Google Scholar
21. 1. Kojima S

    (2015) Dynamism and regulation of the stator, the energy conversion complex of the bacterial flagellar motor

    Current Opinion in Microbiology 28:66–71.

    https://doi.org/10.1016/j.mib.2015.07.015

    • PubMed
    • Google Scholar
22. 1. Kremer JR
    2. Mastronarde DN
    3. McIntosh JR

    (1996) Computer visualization of three-dimensional image data using IMOD

    Journal of Structural Biology 116:71–76.

    https://doi.org/10.1006/jsbi.1996.0013

    • PubMed
    • Google Scholar
23. 1. Kubori T
    2. Shimamoto N
    3. Yamaguchi S
    4. Namba K
    5. Aizawa S

    (1992) Morphological pathway of flagellar assembly in salmonella typhimurium

    Journal of Molecular Biology 226:433–446.

    https://doi.org/10.1016/0022-2836(92)90958-M

    • PubMed
    • Google Scholar
24. 1. Kühn MJ
    2. Schmidt FK
    3. Eckhardt B
    4. Thormann KM

    (2017) Bacteria exploit a polymorphic instability of the flagellar filament to escape from traps

    PNAS 114:6340–6345.

    https://doi.org/10.1073/pnas.1701644114

    • PubMed
    • Google Scholar
25. 1. Macnab RM

    (1999)

    The bacterial flagellum: reversible rotary propellor and type III export apparatus

    Journal of Bacteriology 181:7149–7153.

    • PubMed
    • Google Scholar
26. 1. Magariyama Y
    2. Sugiyama S
    3. Muramoto K
    4. Maekawa Y
    5. Kawagishi I
    6. Imae Y
    7. Kudo S

    (1994) Very fast flagellar rotation

    Nature 371:752.

    https://doi.org/10.1038/371752b0

    • PubMed
    • Google Scholar
27. 1. Minamino T
    2. Imada K

    (2015) The bacterial flagellar motor and its structural diversity

    Trends in Microbiology 23:267–274.

    https://doi.org/10.1016/j.tim.2014.12.011

    • PubMed
    • Google Scholar
28. 1. Morimoto YV
    2. Minamino T

    (2014) Structure and function of the bi-directional bacterial flagellar motor

    Biomolecules 4:217–234.

    https://doi.org/10.3390/biom4010217

    • PubMed
    • Google Scholar
29. 1. Myers CR
    2. Nealson KH

    (1988) Bacterial manganese reduction and growth with manganese oxide as the sole electron acceptor

    Science 240:1319–1321.

    https://doi.org/10.1126/science.240.4857.1319

    • PubMed
    • Google Scholar
30. 1. Nicastro D
    2. Schwartz C
    3. Pierson J
    4. Gaudette R
    5. Porter ME
    6. McIntosh JR

    (2006) The molecular architecture of axonemes revealed by cryoelectron tomography

    Science 313:944–948.

    https://doi.org/10.1126/science.1128618

    • PubMed
    • Google Scholar
31. 1. Oikonomou CM
    2. Jensen GJ

    (2017) A new view into prokaryotic cell biology from electron cryotomography

    Nature Reviews Microbiology 15:128.

    https://doi.org/10.1038/nrmicro.2016.195

    • PubMed
    • Google Scholar
32. 1. Paulick A
    2. Delalez NJ
    3. Brenzinger S
    4. Steel BC
    5. Berry RM
    6. Armitage JP
    7. Thormann KM

    (2015) Dual stator dynamics in the S hewanella oneidensis MR-1 flagellar motor: shewanella oneidensis MR-1 flagellar motor

    Molecular Microbiology 96:993–1001.

    https://doi.org/10.1111/mmi.12984

    • Google Scholar
33. Book

    1. Pfeffer S
    2. Förster F

    (2018) Structural Biology in Situ Using Cryo-Electron Subtomogram Analysis

    In: Hanssen E, editors. Cellular Imaging: Electron Tomography and Related Techniques. Cham: Springer International Publishing. pp. 237–259.

    https://doi.org/10.1007/978-3-319-68997-5_9

    • Google Scholar
34. 1. Pirbadian S
    2. Barchinger SE
    3. Leung KM
    4. Byun HS
    5. Jangir Y
    6. Bouhenni RA
    7. Reed SB
    8. Romine MF
    9. Saffarini DA
    10. Shi L
    11. Gorby YA
    12. Golbeck JH
    13. El-Naggar MY

    (2014) Shewanella oneidensis MR-1 nanowires are outer membrane and periplasmic extensions of the extracellular electron transport components

    PNAS 111:12883–12888.

    https://doi.org/10.1073/pnas.1410551111

    • PubMed
    • Google Scholar
35. 1. Sowa Y
    2. Berry RM

    (2008) Bacterial flagellar motor

    Quarterly Reviews of Biophysics 41:103–132.

    https://doi.org/10.1017/S0033583508004691

    • PubMed
    • Google Scholar
36. 1. Stock D
    2. Namba K
    3. Lee LK

    (2012) Nanorotors and self-assembling macromolecular machines: the torque ring of the bacterial flagellar motor

    Current Opinion in Biotechnology 23:545–554.

    https://doi.org/10.1016/j.copbio.2012.01.008

    • PubMed
    • Google Scholar
37. 1. Subramanian P
    2. Pirbadian S
    3. El-Naggar MY
    4. Jensen GJ

    (2018) Ultrastructure of shewanella oneidensis MR-1 nanowires revealed by electron cryotomography

    PNAS 115:E3246–E3255.

    https://doi.org/10.1073/pnas.1718810115

    • PubMed
    • Google Scholar
38. 1. Terahara N
    2. Krulwich TA
    3. Ito M

    (2008) Mutations alter the sodium versus proton use of a Bacillus clausii flagellar motor and confer dual ion use on Bacillus subtilis motors

    PNAS 105:14359–14364.

    https://doi.org/10.1073/pnas.0802106105

    • PubMed
    • Google Scholar
39. 1. Terahara N
    2. Sano M
    3. Ito M

    (2012) A Bacillus flagellar motor that can use both na+ and K+ as a coupling ion is converted by a single mutation to use only na+

    PLOS ONE 7:e46248.

    https://doi.org/10.1371/journal.pone.0046248

    • PubMed
    • Google Scholar
40. 1. Terashima H
    2. Fukuoka H
    3. Yakushi T
    4. Kojima S
    5. Homma M

    (2006) The Vibrio motor proteins, MotX and MotY, are associated with the basal body of Na ⁺ -driven flagella and required for stator formation

    Molecular Microbiology 62:1170–1180.

    https://doi.org/10.1111/j.1365-2958.2006.05435.x

    • Google Scholar
41. 1. Terashima H
    2. Koike M
    3. Kojima S
    4. Homma M

    (2010) The flagellar basal body-associated protein FlgT is essential for a novel ring structure in the sodium-driven vibrio motor

    Journal of Bacteriology 192:5609–5615.

    https://doi.org/10.1128/JB.00720-10

    • PubMed
    • Google Scholar
42. 1. Terashima H
    2. Li N
    3. Sakuma M
    4. Koike M
    5. Kojima S
    6. Homma M
    7. Imada K

    (2013) Insight into the assembly mechanism in the supramolecular rings of the sodium-driven vibrio flagellar motor from the structure of FlgT

    PNAS 110:6133–6138.

    https://doi.org/10.1073/pnas.1222655110

    • PubMed
    • Google Scholar
43. 1. Terashima H
    2. Kawamoto A
    3. Morimoto YV
    4. Imada K
    5. Minamino T

    (2017) Structural differences in the bacterial flagellar motor among bacterial species

    Biophysics and Physicobiology 14:191–198.

    https://doi.org/10.2142/biophysico.14.0_191

    • PubMed
    • Google Scholar
44. 1. Thormann KM
    2. Paulick A

    (2010) Tuning the flagellar motor

    Microbiology 156:1275–1283.

    https://doi.org/10.1099/mic.0.029595-0

    • PubMed
    • Google Scholar
45. 1. Toutain CM
    2. Caizza NC
    3. Zegans ME
    4. O'Toole GA

    (2007) Roles for flagellar stators in biofilm formation by pseudomonas aeruginosa

    Research in Microbiology 158:471–477.

    https://doi.org/10.1016/j.resmic.2007.04.001

    • PubMed
    • Google Scholar
46. 1. Williams KP
    2. Gillespie JJ
    3. Sobral BW
    4. Nordberg EK
    5. Snyder EE
    6. Shallom JM
    7. Dickerman AW

    (2010) Phylogeny of gammaproteobacteria

    Journal of Bacteriology 192:2305–2314.

    https://doi.org/10.1128/JB.01480-09

    • PubMed
    • Google Scholar
47. 1. Wu L
    2. Wang J
    3. Tang P
    4. Chen H
    5. Gao H

    (2011) Genetic and molecular characterization of flagellar assembly in shewanella oneidensis

    PLOS ONE 6:e21479.

    https://doi.org/10.1371/journal.pone.0021479

    • PubMed
    • Google Scholar
48. 1. Zhao X
    2. Norris SJ
    3. Liu J

    (2014) Molecular architecture of the bacterial flagellar motor in cells

    Biochemistry 53:4323–4333.

    https://doi.org/10.1021/bi500059y

    • PubMed
    • Google Scholar
49. 1. Zheng SQ
    2. Keszthelyi B
    3. Branlund E
    4. Lyle JM
    5. Braunfeld MB
    6. Sedat JW
    7. Agard DA

    (2007) UCSF tomography: an integrated software suite for real-time electron microscopic tomographic data collection, alignment, and reconstruction

    Journal of Structural Biology 157:138–147.

    https://doi.org/10.1016/j.jsb.2006.06.005

    • PubMed
    • Google Scholar
50. 1. Zhu S
    2. Nishikino T
    3. Hu B
    4. Kojima S
    5. Homma M
    6. Liu J

    (2017) Molecular architecture of the sheathed polar flagellum in vibrio alginolyticus

    PNAS 114:10966–10971.

    https://doi.org/10.1073/pnas.1712489114

    • PubMed
    • Google Scholar

Author details

1. Mohammed Kaplan

   Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, United States

   Contribution

   Conceptualization, Data curation, Formal analysis, Funding acquisition, Validation, Investigation, Visualization, Methodology, Writing—original draft, Writing—review and editing

   Competing interests

   No competing interests declared

   "This ORCID iD identifies the author of this article:" 0000-0002-0759-0459

2. Debnath Ghosal

   Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, United States

   Contribution

   Data curation, Writing—review and editing

   Competing interests

   No competing interests declared

3. Poorna Subramanian

   Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, United States

   Contribution

   Data curation, Writing—review and editing

   Competing interests

   No competing interests declared

4. Catherine M Oikonomou

   Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, United States

   Contribution

   Formal analysis, Writing—review and editing

   Competing interests

   No competing interests declared

5. Andreas Kjaer

   Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, United States

   Present address

   University of Oxford, Oxford, United Kingdom

   Contribution

   Formal analysis, Writing—review and editing

   Competing interests

   No competing interests declared

   "This ORCID iD identifies the author of this article:" 0000-0002-0096-5764

6. Sahand Pirbadian

   Department of Physics and Astronomy, Biological Sciences, and Chemistry, University of Southern California, Los Angeles, United States

   Contribution

   Methodology, Writing—review and editing

   Competing interests

   No competing interests declared

7. Davi R Ortega

   Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, United States

   Contribution

   Methodology, Writing—review and editing

   Competing interests

   No competing interests declared

8. Ariane Briegel

   Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, United States

   Present address

   Institute of Biology, Leiden University, Leiden, The Netherlands

   Contribution

   Data curation, Writing—review and editing

   Competing interests

   No competing interests declared

   "This ORCID iD identifies the author of this article:" 0000-0003-3733-3725

9. Mohamed Y El-Naggar

   Department of Physics and Astronomy, Biological Sciences, and Chemistry, University of Southern California, Los Angeles, United States

   Contribution

   Resources, Writing—review and editing

   Competing interests

   No competing interests declared

10. Grant J Jensen

    1. Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, United States
    2. Howard Hughes Medical Institute, California Institute of Technology, Pasadena, United States

    Contribution

    Conceptualization, Resources, Formal analysis, Supervision, Funding acquisition, Validation, Investigation, Project administration, Writing—review and editing

    For correspondence

    jensen@caltech.edu

    Competing interests

    No competing interests declared

    "This ORCID iD identifies the author of this article:" 0000-0003-1556-4864

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