1.A.30 The H+- or Na+-translocating Bacterial Flagellar Motor/ExbBD Outer Membrane Transport Energizer (Mot-Exb) Superfamily
The Mot-Exb Superfamily consists of six distant families, each probably with a distinct physiological function, although all may function as H+/Na+ channels, driving an energy-requiring process. For example, flagellar motors of marine bacteria Halomonas are driven by both protons and sodium ions (Kita-Tsukamoto et al. 2004). The MotAB family energizes bacterial flagellar rotation while the ExbBD family energizes accumulation of large molecules (i.e. iron-siderophores, vitamin B12, DNA from phage, and colicins) from the external medium across the outer Gram-negative bacterial membrane into the periplasm. The AglRS system powers gliding motility while the SilAB systems energize gian adhesin export. The function of a 5th family (TC# 1.A.30.5) is not known, but the ZorAB systems have been reported to function as parts of antiphage defense systems. The pmf (or smf) is probably the driving force in all cases. MotAB and PomAB are homologous to ExbBD and TolQR. MotAB of E. coli, the stator, is known to form a proton channel. This stator is composed of MotA and MotB proteins, which form a hetero-hexameric complex with a stoichiometry of four MotA and two MotB molecules. MotA can form a tetramer in the absence of MotB (Takekawa et al. 2016). Ion binding residues for Na+ flow in the stator complex of the Vibrio flagellar motor have been identified (Onoue et al. 2019). The structure and dynamics of the bacterial flagellum have been reviewed (Nakamura and Minamino 2019). The flagellar motor, which structurally resembles an artificial motor, is embedded within the cell envelop and spins at several hundred revolutions per second (Morimoto and Minamino 2021). A single nucleotide polymorphism alters the activity of the renal Na+:Cl- cotransporter and reveals a role for transmembrane segment 4 in chloride and thiazide affinity (Moreno et al. 2004).
About 10 stators (MotA/MotB complexes) are docked around a rotor, and the stator recruitment depends on the load, ion motive force, and coupling ion flux. The MotA(M206I) mutation slows motor rotation and decreases the number of docked stators in Salmonella. Suzuki et al. 2019 showed that lowering the external pH improves the assembly of the mutant stators. Neither the collapse of the ion motive force nor a mutation mimicking the proton-binding state inhibited stator localization to the motor. Thus, MotA-Met206 is involved in torque generation and proton translocation, and stator assembly is stabilized by protonation of the stator. Ancestral sequence reconstructions of MotB require MotA and give rise to pmf-dependent motility (Islam et al. 2020).
Yonekura et al. (2011) presented the first three-dimensional structure of the PomAB torque-generating stator unit analyzed by electron microscopy. The structure of PomAB revealed two arm domains, which contain the PG-binding site, connected to a large base made of the transmembrane and cytoplasmic domains. The arms lean downward to the membrane surface, likely representing a 'plugged' conformation, which would prevent ions leaking through the channel. They propose a model for how PomAB units are placed around the flagellar basal body to function as torque generators.
Leu46 of MotB acts as the gate for hydronium ion permeation, which induces the formation of a water wire that may mediate the proton transfer to Asp32 on MotB. The free energy barrier for H3O+ permeation is consistent with the proton transfer rate deduced from the flagellar rotational speed and the number of protons per rotation, suggesting that gating is the rate-limiting step (Nishihara and Kitao 2015). Structure and dynamics of MotA/B with nonprotonated and protonated Asp32 suggested size-dependent ion selectivity. In MotA/B with the nonprotonated Asp32, the A3 segment in MotA maintains a kink whereas protonation induces a straighter shape. Assuming that the cytoplasmic domain not included in the atomic model moves as a rigid body, the protonation/deprotonation of Asp32 is inferred to induce a ratchet motion of the cytoplasmic domain, correlated with the motion of the flagellar rotor (Nishihara and Kitao 2015).
ExbBD forms both hexameric and pentameric complexes that coexist, with the proportion of the hexamers increasing with pH. Channel current measurements and 2D crystallography thus support the existence of and transition between the two oligomeric states in membranes. The hexameric complex has been reported to consist of six ExbB subunits and three ExbD transmembrane helices enclosed within the central channel (Maki-Yonekura et al. 2018). TonB physically interacts with the nutrient-loaded transporter to exert a force that opens an import pathway across the outer membrane. Another group showed that five copies of ExbB are arranged as a pentamer around two copies of ExbD in the complex. The revised stoichiometry has implications for motor function (Celia et al. 2019).
As noted above, each flagellum is a supramolecular motility machine consisting of a bi-directional rotary motor, a universal joint and a helical propeller. The signal transducers transmit environmental signals to the flagellar motor through the cytoplasmic chemotactic signaling pathway. The flagellar motor is composed of a rotor and multiple stator units, each of which acts as a transmembrane proton channel to conduct protons and exert force on the rotor (Minamino et al. 2019). FliG, FliM and FliN form the C ring on the cytoplasmic face of the basal body MS ring made of the transmembrane protein FliF and act as the rotor. The C ring also serves as a switching device that enables the motor to spin in both counterclockwise (CCW) and clockwise (CW) directions. The phosphorylated form of the chemotactic signaling protein CheY binds to FliM and FliN to induce conformational changes of the C ring responsible for switching the direction of flagellar motor rotation from CCW to CW (Minamino et al. 2019).
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).
The flagellar motor (pmf-dependent) (MotA-MotB). TMSs 3 and 4 of MotA and the single TMS of MotB comprise the proton channel, which is inactive until the complex assembles into a motor. Hosking et al. 2006 identify a periplasmic segment of the MotB protein that acts as a plug to prevent premature proton flow. The plug is in the periplasm just C-terminal to the MotB TMS flanked by Pro52 and Pro65. The Pro residues and Ile58, Tyr61, and Phe62 are essential for plug function (Hosking et al. 2006).
The mechanism of proton passage and coupling to flagellar rotation has been proposed (Nishihara and Kitao 2015). About a dozen MotA/B complexes are anchored to the peptidoglycan layer around the motor through the C-terminal peptidoglycan-binding domain of MotB (Castillo et al. 2013). Dynamic permeation by hydronium ions, sodium ions, and water molecules has been observed using steered molecular dynamics simulations, and free energy profiles for ion/water permeation were calculated (Kitao and Nishihara 2017). They also examined the possible ratchet motion of the cytoplasmic domain induced by the protonation/deprotonation cycle of the MotB proton binding site, Asp32. The motor (MotAB) consists of a dynamic population of mechanosensitive stators that are embedded in the inner membrane and activate in response to external load. This entails assembly around the rotor, anchoring to the peptidoglycan layer to counteract torque from the rotor and opening of a cation channel to facilitate an influx of cations, which is converted into mechanical rotation. Stator complexes are comprised of four copies of an integral membrane A subunit and two copies of a B subunit. Each B subunit includes a C-terminal OmpA-like peptidoglycan-binding (PGB) domain. This is thought to be linked to a single N-terminal transmembrane helix by a long unstructured peptide, which allows the PGB domain to bind to the peptidoglycan layer during stator anchoring. The high-resolution crystal structures of flagellar motor PGB domains from Salmonella enterica have been solved (Liew et al. 2017). Change in the C ring conformation for switching and rotation involve loose and tight intersubunit interactions (Sakai et al. 2019).
MotA and MotB of E. coli
The flagellar motor (smf-dependent) (PomAB; MotXY) (Okabe et al., 2005). PomB interacts with the third TMS of PomA in the Na+-driven polar flagellum (Yakushi et al. 2004). Sodium-powered stators of the flagellar motor can generate torque in the presence of the sodium channel blocker, phenamil, with mutations near the peptidoglycan-binding region of PomB (Ishida et al. 2019). FliL associates with the flagellar stator in the sodium-driven Vibrio motor (Lin et al. 2018). When the ion channel is closed, PomA and PomB interact strongly. When the ion channel opens, PomA interacts less tightly with PomB. The plug and loop between TMSs 1 and 2 regulate activation of the stator, which depends on the binding of sodium ion to the D24 residue of PomB (Nishikino et al. 2019). The PomA helices parallel to the inner membrane play iroles in the hoop-like function in securing the stability of the stator complex and the ion conduction pathway (Nishikino et al. 2022).
PomAB/MotXY of Vibrio alginolyticus
The flagellar motor (pmf-dependent) (MotAB) (Ito et al., 2004)
MotAB of Bacillus subtilis
The flagellar motor (smf-dependent) (MotPS) (Ito et al., 2004)
MotPS (YtxDE) of Bacillus subtilis
MotP (YtxD) (MotA homologue) (P39063)
MotS (YtxE) (MotB homologue) (P39064)
The H+-driven flagellar motor complex, MotABXY (MotXY are required for systems 1.A.30.1.5 and 1.A.30.1.6; Koerdt et al., 2009).
The H+-driven flagellar motor complex of Shewanella oneidensis
The Na+-driven flagellar motor complex, PomAB MotXY (MotXY are required for systems 1.A.30.1.5 and 1.A.30.1.6; Koerdt et al., 2009)
The Na+-driven flagellar motor complex of Shewanella oneidensis
The motor complex of the bacterial flagellum, MotAB. MotA is 295 aas long with about 5 putative TMSs in a 2 + 1 + 2 TMS arrangement, possibly with a C-terminal additional TMS. MotB is 309 aas long with a single N-terminal TMS. They comprise the stator element of the flagellar motor
complex and are required for rotation of the flagellar motor. Together they form the transmembrane proton channel. These two proteins are 94 and 91% identical to the E. coli complex (TC# 1.A.30.1.1) (Morimoto and Minamino 2014).
MotAB of Salmonella enterica, subspecies Typhimurium
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).
The TonB energy-transducing system. ExbB/D (the putative H+ channel) are listed here; TonB is listed under TC# 2.C.1.1.1. Deletion of the cytoplasmic loop gives rise to immediate growth arrest (Bulathsinghala et al. 2013). The rotational surveillance and energy transfer (ROSET) model of TonB action postulates a mechanism for the transfer of energy from the IM to the OM, triggering iron uptake and concentration in the periplasm (Klebba 2016).
The TonB system of E. coli
Putative biopolymer transport proteins ExbB/ExbD-like 3 (Sll1404/Sll1405). Involved in the TonB-dependent energy-dependent transport of iron-siderophores via FhuA (Sll1406; TC# 1.B.14.2.9) using TonB (TC# 2.C.1.3.1) (Qiu et al. 2018). ExbB may protect ExbD from proteolytic degradation and functionally stabilizes TonB.
ExbBD of Synechocystis sp. (strain PCC 6803 / Kazusa)
ExbB/ExbD/ExbD' of 254 aas, 150 aas, and 147 aas, respectively, with 3, 1 and 1 TMSs, respectively. The genes encoding these three proteins are adjacent to each other. ExbD and TonB (TC# 2.C.1.3.1) interact directly (Qiu et al. 2018).
ExbB/ExbD/ExbD' of Synechocystis sp. (strain PCC 6803 / Kazusa)
The TolA energy-transducing system. TolQ/R (the putative H+ channel) are listed here; TolA is listed under TC# 2.C.1.2.1, together with its auxiliary proteins. The channel is lined by TolR-Asp23, TolQ-Thr145 and TolQ-Thr178. The Tol-Pal complex, energized by TolQRA, and using the outer membrane proteins, BtuB and OmpF as receptors, is responsible for the uptake of colicin ColE9 and other bacteriocins; in this process, the complex in the outer membrane bridges and immobilizes the complex components in the inner membrane (Rassam et al. 2018). Salmonella Tol-Pal reduces outer membrane glycerophospholipid levels for envelope Hhomeostasis and survival during bacteremia in a process dependent on the TolQR channel (Masilamani et al. 2018).
The TolA system of E. coli
Putative TolA Energizer, TolQ1/TolR1
TolQ1/R1 of Myxococcus xanthus
Putative TolA Energizer, TolQ2/TolR2
TolQ2/R2 of Myxococcus xanthus
Putative TolA Energizer, TolQ3/TolR3
TolQ3/R3 of Myxococcus xanthus
Putative TolA Energizer, TolQ4/TolR4
TolQ4/R4 of Myxococcus xanthus
Putative TolA-dependent Energizer, TolQ5/TolR5 or AglX/AglV. Identified as an essential motor for adventurous gliding motility (Nan et al. 2011).
TolQ5/R5 or AglX/AglV of Myxococcus xanthus
The putative ExbBD energizer (H+-channel).
ExbBD of Leptospira interrogans
TolQ/R of Leptospira interrogans
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).
TolQ/TolR of Geobacter sp. M18
Motor for adventurous motility, AglR (a TolQ homologue)/AglS (a TolR homologue) (Nan et al. 2011). The mechanism by which the AglRS proteins energize adventurous gliding motility has been proposed (Jakobczak et al. 2015; Mignot and Nöllmann 2017).
AglQ/AglR of Myxococcus xanthus
Adventurous gliding motility proteins AglR, AglS and AglV. These three proteins presumably function in gliding motility but are homologues of MotA/ExbB, MotB/ExbD and MotB/ExbD, respectively.
AglRSV of Bdellovibrio bacteriovorus
AglR, 248 aas and 3 TMSs (Q6MQP0)
AglS, 165 aas and 1 TMS (Q6MQP2)
AglV, 149 aas and 1 TMS (Q6MQP1)
Putative gliding motility energizing system, AglR/AglS/AglS'. Similar to the three components of another system in the same organism with TC# 1.A.30.3.3.
AglR/S/S' of Bdellovibrio bacteriovorus
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.
SiiAB putative energizer of giant adhesin, SiiE (repetitive 5,559 aa protein) export (Wille et al. 2013).
SiiAB of Salmonella enterica
SiiA (210 aas; 1 TMS) (E1WEU7)
SiiB (462 aas; 3 TMSs) (E1WEU8)
SiiA/B (MotB/A) homologues of 236 and 362 aas with 1 and 4 putative TMSs, respectively.
SiiA/B of Sulfurimonas denitrificans (Thiomicrospira denitrificans)
SiiA (MotB) homologue of 236 aas (Q30RA9)
SilB (MotA) homologue of 362 aas (Q30RB0)
SiiA/B (MotB/A) homologues of 315 and 350 aas with 1 and 4 putative TMSs, respectively. Functions with the Type I (ABC) protein secretion exporter of TC# 3.A.1.109.6).
SiiA/B homologues of Desulfovibrio salexigens
SiiA homologue (315 aas; C6BWI8)
SiiB homologue (350 aas; C6BWI5)
Uncharacterized MotA homologue of 301 aas and 3 to 5 TMSs. If 5, TMSs 1, 4 and 5 are strongly hydrophobic while TMSs2 and 3 are quite hydrophilic. The gene encoding this protein is flanked by genes encoding two large proteins, one anotated as a microtubule binding protein but showing limited sequence similarity to TolA (522 aas and one N-terminal TMS; Q6MNS1) and the other a large Ala/Glu-rich protein (794 aas with an N-terminal phosphopeptide binding motif (FHA), two ATP synthase domains (B and E), and one TMS near the C-terminus; Q6MNS3).
UP of Bdellovibrio bacteriovorus
ZorA/ZorB forming a putative proton channel. ZorA (704 aas; 4 TMSs; ABS66238; a putative methyl chemotaxis protein) is a very distant MotA homologue showing most sequence similarity to TC# 1.A.30.5.1) while ZorB is a clear MotB homologue (254 aas with 1 N-terminal TMS; ABS66239; hits 1.A.30.1.4 with a score of e-09). This system is proposed to function as an anti-phage defense system. ZorAB are only two such components, suggested to have proton channel activity (Doron et al. 2018).
ZorAB of Xanthobacter autotrophicus Py2
ZorA/ZorB (MotAB homologues) of 704 aas with 3 TMSs and 254 aas with 1 N-terminal TMS.
ZorAB of Pseudomonas aeruginosa
ZorA/ZorB components of an anti-phage defense system (Doron et al. 2018).
ZorAB of Acinetobacter baumannii
ZorAB, putative H+ channel proteins, MotAB homologues, functioning in defense against phage attack (Doron et al. 2018).
ZorAB of E. coli PA10
ZorAB, putative proton channel proteins, homologues of MotAB, possibly involved in resistance to phage invasion (Doron et al. 2018).
ZorAB of Acidaminococcus fermentans DSM 20731
ZorA (673 aas amd 3 - 4 TMSs)/ZorB (248 aas and 1 N-terminal TMS)
ZorAB of Prochlorothrix hollandica PCC 9006
ZorAB of Thermosipho africanus
ZorA (472 aas and 3 TMSs)/ZorB (222 aas and 1 N-terminal TMS.
ZorAB of Spirosoma linguale