The flagellar motor complexes of bacteria are driven either by the proton motive force (pmf) or the sodium motive force (smf). The motor in E. coli consists of MotA and MotB, which comprise the stator. MotA interacts with FliG, a cytoplasmic component of the rotor, via electrostatic interactions. The 3-dimensional structure of the C-terminal domain of FliG, which interacts with the motor, has been determined revealing a cluster of charged residues in a ridge that interact with MotA. Additionally, MotA contains a conserved C-terminal cluster of negatively charged residues, and MotB contains a conserved N-terminal cluster of positively charged residues. Hosking & Manson (2008) have provided evidence that these patches of charged residues provide the non-covalent link between the two components of the stator. The MotB linker plays a role in the assembly and activation of the flagellar motor (O'Neill et al., 2011).

Four proteins (PomA, PomB, MotX and MotY) have been implicated in Na⁺-dependent flagellar rotation in Vibrio alginolyticus PomA and PomB are homologous to MotA and MotB, respectively, and they form a PomA:PomB = 2:1 heterotrimeric complex. MotX and MotY have been sequenced from Vibrio species. MotX contains a single putative N-terminal transmembrane spanner (TMS) and a region that exhibits three tandem repeat sequences, each 36 amino acyl residues in length, with similarity to repeat sequences in two ORFs from E. coli (e.g., acc# gbU82598) and in an ORF from Helicobacter pylori (gbU86608). MotY possesses a single N-terminal TMS and contains a C-terminal region that exhibits sequence similarity with members of the OmpA-OmpF Porin (OOP) Family (TC #1.B.6) and peptidoglycan-associated lipoproteins. The functional relationship of PomAB to MotXY is not known, but PomAB has been reported to alone be the functional component of the torque-generating unit, and MotX and MotY are not specifically required for smf-coupled flagellar rotation. Purified PomA/PomB reconstituted into proteoliposomes catalyzed ²²Na⁺ influx in response to a K⁺ diffusion potential showing that PomA/B comprise the Na⁺ channel.

PomA and PomB form the Na⁺ channel that functions as a stator complex to couple sodium-ion flux with torque generation. MotX and MotY are components of the T-ring, which is located beneath the P-ring of the polar flagellar basal body and is involved in incorporation of the PomA/PomB complex into the motor. Kojima et al. (2008) described the crystal structure of MotY at 2.9 A resolution. It has two domains: an N-terminal domain (MotY-N) and a C-terminal domain (MotY-C). MotY-N has a unique structure. MotY-C contains a putative peptidoglycan-binding motif that is similar to those of peptidoglycan-binding proteins, but this region is disordered in MotY. Motility assay of cells producing either of the MotY-N and MotY-C fragments and subsequent biochemical analyses indicate that MotY-N is essential for association of the stator units around the rotor, whereas MotY-C stabilizes the association by binding to the peptidoglycan layer.

Phylogenies of the Mot family proteins do not follow the phylogenies of the source organisms suggesting the presence in bacteria of isoforms. MotB of E. coli, which spans the cytoplasmic membrane once, possesses a C-terminal peptidoglycan-interaction domain and probably functions as part of the 'stator' by anchoring the flagellar complex to the cell wall. MotA, which exhibits 4 TMSs, probably comprises most of the ion-translocating 'channel' together with the membrane-embedded helix of MotB. The fully conserved aspartyl residue in the center of the MotB TMS may serve as a cation binding residue. Homologous pairs of MotA - MotB paralogues in a single bacterial species, for example, Bacillus subtilis, are known to exist, where one pair functions in motility while the other pair is of unknown function. The flux of H⁺ or Na⁺ through the stator drives rotation of the rotor which corresponds to the rotating part of the flagellum. MotA-B are homologous to (but distantly related to) ExbB-D (see below).

The Mot complex is anchored to the peptidoglycan (PG) layer by the peptidoglycan-binding (PGB) domain of MotB. Proton translocation is activated only when the stator is installed into the motor. Kojima et al. (2009) reported the crystal structure of a C-terminal periplasmic fragment of MotB (MotB(C)) that contains the PGB domain and includes the entire periplasmic region essential for motility. Structural and functional analyses indicated that the PGB domains must dimerize in order to form the proton-conducting channel. Drastic conformational changes in the N-terminal portion of MotB(C) appear to be required both for PG binding and the proton channel activation (Kojima et al., 2009).

The torque of bacterial flagellar motors is generated by interactions between the rotor and the stator, and is coupled to the influx of H⁺ or Na⁺ through the stator. A chimeric protein PotB, in which the N-terminal region of Vibrio alginolyticus PomB is fused to the C-terminal region of E. coli MotB, can function with PomA as a Na⁺-driven stator in E. coli. Zhu et al. (2012) constructed a deletion variant of PotB (Δ41-91; called PotBΔL), which lacks the periplasmic linker region including the segment that works as a 'plug' to inhibit premature ion influx. This variant did not confer motile ability, but a Na⁺-driven, spontaneous suppressor mutant has a point mutation (R109P) in the MotB/PomB specific α-helix that connects the transmembrane and peptidoglycan binding domains of PotBΔL in the region of MotB and did confer motility. Over-production of the PomA/PotBΔL(R109P) stator inhibited the growth of E. coli cells, suggesting that this stator has high Na⁺-conducting activity. Mutational analyses of Arg109 and nearby residues suggested that the structural alteration in this α-helix optimized the PotBΔL conformation and restored the proper arrangement of transmembrane helices to form a functional channel pore. Possibly, this α-helix plays a key role in assembly-coupled stator activation.

The bacterial flagellar motor consists of an approximately 50-nm rotor with up to 10 independent stators anchored to the cell wall. Lo et al. 2013 measured torque-speed relationships of single-stator motors under 25 different combinations of electrical and chemical potential. All 25 torque-speed curves had the same concave-down shape as fully energized wild-type motors, and each stator passes at least 37 +/- 2 ions per revolution. The motor mechanism has a 'powerstroke' in either ion binding or transit; ion transit is channel-like rather than carrier-like; and the rate-limiting step in the motor cycle is ion binding at low concentration, ion transit, or release at high concentration. 

The flagellar ion channel complex consists of at least three structural parts: a cytoplasmic domain responsible for the interaction with the rotor, a transmembrane ion channel, and a peptidoglycan-binding (PGB) domain. A flexible linker connecting the ion channel and the PGB domain not only coordinates stator assembly with its ion channel activity but also controls the assembly of stator units to the motor in response to changes in the environment. When the ion channel complex encounters the rotor, the N-terminal portion of the PGB domain adopts a partially stretched conformation, allowing the PGB domain to reach and bind to the PG layer. The binding affinity of the PGB domain for the PG layer is affected by the force applied to its anchoring point and to the type of ionic energy source. Minamino et al. 2018 reviewed these aspects of the flagellum and presented a current (2018) understanding of autonomous control mechanism of stator assembly in the bacterial flagellar motor.

Three ~3 Å-resolution cryoEM reconstructions of the stator unit (MotAB) in different functional states have been published (Santiveri et al. 2020). The stator unit consists of a dimer of MotB surrounded by a pentamer of MotA [MotA₅.MotB₂]. Combining structural data with mutagenesis and functional studies, key residues involved in torque generation were determined, and a detailed mechanistic model for motor function and switching of rotational direction was presented (Santiveri et al. 2020).

The generalized transport reaction catalyzed by the MotAB H⁺ channel is:

H⁺ (out) ↔⁺ (in).

That for the Na⁺-driven PomAB motor is:

Na⁺ (out) ↔⁺ (in).

TonB heterotrimeric complexes span the cytoplasmic membrane and the periplasm and interact with outer membrane receptors in Gram-negative bacteria. Homologues have been found only in Gram-negative bacteria and cyanobacteria. E. coli possesses two paralogous systems, the TonB-ExbB-ExbD system and the TolA-TolQ-TolR system. Corresponding proteins have been identified in other Gram-negative bacteria. TonB (239 aas) and TolA (412 aas) of E. coli both span the cytoplasmic membrane once near their N-termini, span the periplasm as α-helices and interact with the outer membrane. They are not demonstrably homologous, but they are believed to serve comparable functions. It has been suggested that TonB or its C-terminal domain shuttles between the cytoplasmic and outer membranes as part of the energy transduction process. ExbB (244 aas) and TolQ (230 aas) are paralogous. They span the membrane 3 times with their N-termini in the periplasm and most of the protein mass localized to the cytoplasmic side of the membrane. ExbD (141 aas) and TolR (142 aas) are also paralogous. They span the membrane near their N-termini with remaining parts of the proteins in the periplasm. The TonB system energizes transport (uptake) via OMR-type porins (TC #1.B.14) of vitamin B₁₂, iron-siderophores, group B colicins and the DNA of filamentous bacteriophage such as φ80 and T1. The TolA system transports group A colicins and the DNA of other filamentous phages. Colicin import requires close proximity of the inner and outer membranes. Loss of one of the TolA-TolQ-TolR proteins results in loss of periplasmic enzymes and increased sensitivity to drugs and bile salts. Surface localization of O-antigen lipopolysaccharide in E. coli depends on the TolA protein, possibly explaining the leakiness of TolA mutants.

ExbB-ExbD are homologous to (but distantly related to) the MotA-MotB proteins. While the MotA-B proteins have been shown to provide a transmembrane proton translocation pathway, the same has not yet been demonstrated for the ExbB-D proteins. It is expected, however, that they will serve this function and thereby energize outer membrane transport. All of these proteins can be considered both as proton channel proteins and as pmf-dependent energizers. They belong in a single family.

Braun and Herrmann (2004) have proposed that at least three well-conserved transmembrane residues in ExbB (or TolQ) comprise the proton pathway. These proposed channel residues in ExbB are Thr148 in TMS2 and glu176 and Thr181 in TMS3. The first two are strictly conserved in all ExbB and TolQ homologues, and the third is almost strictly conserved in all MotA homologues as well. Asp25 in ExbD may also comprise part of this proton pathway (Braun and Herrmann, 2004). A PMF-dependent TonB-ExbD interaction is prevented by 10-residue deletions within a periplasmic disordered domain of ExbD adjacent to the cytoplasmic membrane. A conserved motif, (V45, V47, L49, P50), within the disordered domainof ExbD is required for signal transduction to TonB. This conserved motif transduces signals to distal regions of ExbD, and then to TonB (Kopp and Postle 2020).

ExbD forms complexes with TonB, ExbB and itself (homodimers) in vivo. The pmf was required for detectable cross-linking between the TonB-ExbD periplasmic domains (Ollis et al., 2009). The presence of inactivating transmembrane domain mutations ExbD(D25N) or TonB(H20A) prevented efficient formaldehyde cross-linking between ExbD and TonB. Neither TonB, ExbB nor the pmf was required for ExbD dimer formation. Two models were considered where either dynamic complex formation occurred through transmembrane domains or the transmembrane domains of ExbD and TonB configure their respective periplasmic domains.

The generalized transport process energized by TonB-type systems is:

substrate (out) substrate (periplasm).

Myxococcus xanthus glides over surfaces using two motility systems social-motility, powered by the retraction of type IV pili, and adventurous (A)-motility, powered by unknown mechanism(s). Nan et al. (2011) showed that AgmU, an A-motility protein, is part of a multiprotein complex that spans the inner membrane and periplasm of M. xanthus and presented evidence that periplasmic AgmU decorates a looped continuous helix that rotates clockwise as cells glide forward, reversing its rotation when cells reverse polarity. Inhibitor studies showed that the AgmU helix rotation is driven by the proton motive force and depends on actin-like MreB cytoskeletal filaments. The AgmU motility complex interacts with the MotAB homologs, AglQR, the presumed channel-forming motor. A mechanochemical model was suggested in which driven motors, similar to bacterial flagellar stator complexes, run along an endless looped helical track, driving rotation of the track. Deformation of the cell surface by the AgmU-associated proteins may create pressure waves in the slime, pushing cells forward (Nan et al. 2011).

The giant non-fimbrial adhesin SiiE of 5559 aas is essential to establish intimate contact between Salmonella enterica and the apical surface of polarized epithelial cells. SiiE is secreted by a type I secretion system (T1SS) encoded by Salmonella Pathogenicity Island 4 (SPI4). Wille et al. (2013) identified SiiA and SiiB as two regulatory proteins encoded by SPI4. Mutant strains in siiA or siiB still secrete SiiE, but are highly reduced in adhesion to and invasion of polarized cells. SiiA and SiiB are inner membrane proteins with one and three transmembrane (TM) helices, respectively. TMS2 and TMS3 of SiiB are similar to members of the ExbB/TolQ family, while the TMS of SiiA is similar to that of MotB, and a conserved aspartyl residue in this TMS is essential for SPI4-encoded T1SS function. Co-immunoprecipitation, bacterial two hybrid and FRET techniques demonstrated homo- and heterotypic protein interactions for SiiA and SiiB. SiiB, but not SiiA also interacts with the SPI4-T1SS ABC-type integral membrane-ATPase, SiiF, which probably exports the giant, repetitive, non-fibrial adhesin of 5,559 aas, SiiE, possibly with the aid of the pmf, using SiiAB as the energizer. The integrity of the Walker A box in SiiF was required for SiiB-SiiF interaction and SiiF dimer formation. SiiA and SiiB appear to be novel virulence-associated members of the Mot/Exb/Tol family of membrane proteins. These two proteins are involved in a mechanism of controlling SPI4-T1SS-dependent adhesion, most likely by formation of a proton-conducting channel and partially energizing SiiE export.  Possibly, ATP hydrolysis by SiiF initiates export, and SiiAB and the pmf energize continued export of the huge SiiE protein.