The research report should be a detailed narrative explaining the function, biological processes, and localization of the gene product. Citations should be given for all claims.
You should prioritize authoritative reviews and primary scientific literature when conducting research. You can supplement
this with annotations you find in gene/protein databases, but these can be outdated or inaccurate.
We are specifically interested in the primary function of the gene - for enzymes, what reaction is catalyzed, and what is the substrate specificity? For transporters, what is the substrate? For structural proteins or adapters, what is the broader structural role? For signaling molecules, what is the role in the pathway.
We are interested in where in or outside the cell the gene product carries out its function.
We are also interested in the signaling or biochemical pathways in which the gene functions. We are less interested in broad pleiotropic effects, except where these elucidate the precise role.
Include evidence where possible. We are interested in both experimental evidence as well as inference from structure, evolution, or bioinformatic analysis. Precise studies should be prioritized over high-throughput, where available.
The requested protein is FliI, the cytoplasmic ATPase of the flagellar type III secretion system (fT3SS) in Salmonella enterica serovar Typhimurium LT2 (ordered locus STM1972; historical synonyms flaAIII and flaC). Its primary biochemical activity is ATP hydrolysis, conventionally written ATP + H₂O → ADP + Pi. Functionally, however, FliI is best understood not as a continuously operating membrane transporter, but as a dynamic substrate-delivery and export-gate-activation ATPase that enables efficient, correctly ordered construction of the flagellar rod, hook, junction, filament, and cap.
FliI forms a six-membered ATPase ring beneath the flagellar export gate. Together with 12 FliH molecules and one central FliJ molecule, it produces an FliH₁₂–FliI₆–FliJ₁ complex analogous in overall architecture to the soluble sector of F₀F₁ ATP synthase. ATP-dependent conformational changes activate or reset the membrane export machinery, whereas the principal energy for processive polypeptide translocation is normally supplied by proton motive force (PMF). FliI therefore increases export efficiency, substrate delivery, and fidelity, but ATP hydrolysis is neither obligatorily coupled one-for-one to exported subunits nor the sole energy source for transport. (minamino2022insightintodistinct pages 1-2, minamino2014thebacterialflagellar pages 1-2)
The supplied identity is internally consistent:
The literature retrieved for Salmonella FliI consistently describes a soluble/cytoplasmic flagellar-export ATPase that forms a homohexamer, interacts with FliH and FliJ, docks beneath the FlhA/FlhB export platform, and supports flagellar protein export. These properties align with the supplied protein description and domains. (kinoshita2021apositivecharge pages 1-2, minamino2022insightintodistinct pages 1-2)
Uppercase human FLII denotes “FLII actin remodeling protein,” a eukaryotic protein associated in Open Targets with human disease phenotypes; it is unrelated to bacterial flagellar FliI. Those human associations must not be transferred to P26465. (OpenTargets Search: -fliI)
The evidence matrix below summarizes the annotation.
| Feature | Best-supported annotation | Evidence type | Key quantitative detail |
|---|---|---|---|
| Identity | FliI (UniProt P26465; locus STM1972) is the flagellum-specific type III protein-export ATPase of Salmonella enterica serovar Typhimurium LT2, not the unrelated human FLII actin-remodeling protein. | Curated annotation, sequence-family assignment, and Salmonella genetic/biochemical studies | Approximately 49 kDa; belongs to the ATPase α/β-chain-related T3SS ATPase family |
| Enzymatic reaction | Catalyzes ATP hydrolysis: ATP + H₂O → ADP + phosphate. ATP is the experimentally supported nucleotide substrate; convincing physiological activity toward alternative nucleotides has not been established. | Purified-protein ATPase assays and catalytic-mutant analysis | Six catalytic sites form at FliI–FliI subunit interfaces in the assembled ring |
| Cellular localization | Soluble cytoplasmic/peripheral-membrane protein that partitions between a freely diffusing pool and the cytoplasmic face of the flagellar basal body; it is not an integral membrane transporter. | Fluorescence localization, turnover measurements, and interaction studies | Approximately 90% of observed basal-body FliI fluorescent spots exchanged with the cytoplasmic pool |
| Oligomer and complex | FliI assembles as a homohexamer around the central-stalk protein FliJ; six FliH dimers bind the FliI N-terminal domains and anchor the ATPase beneath the export gate. Cytoplasmic FliI also occurs in a mobile FliH₂–FliI heterotrimer. | Structural, biochemical, imaging, and review-level synthesis | Basal complex stoichiometry: FliH₁₂–FliI₆–FliJ₁; FliI ring approximately 10 nm across with a roughly 2.5–3.0 nm central cavity |
| Major partners | FliH regulates FliI and connects it to FliN/flagellar C-ring components; FliJ occupies the FliI pore and contacts FlhA; FlhA and FlhB form the cytoplasmic docking platform. FliI-containing carriers also associate with export substrates and chaperone–substrate complexes. | Pull-down, genetic interaction, structural, and dynamic-localization evidence | Export gate includes a FlhA nonamer plus FlhB and the FliP₅–FliQ₄–FliR channel complex |
| Mechanistic role | Promotes substrate delivery to the export gate and uses ATP binding/hydrolysis-driven conformational changes to activate or reset the ion-motive-force-driven export machinery. It increases efficiency and fidelity but is not the membrane translocation pore or the principal continuous energy source. | Mutant, suppressor, secretion, motility, and energy-coupling experiments | Activated gate functions primarily as an H⁺/protein antiporter; T3SS transport can reach tens of thousands of amino acids per second |
| Catalytic-mutant evidence | Glu211 substitutions separate ATP binding/hydrolysis from ongoing protein transport: E211D permits very infrequent hydrolysis yet substantial export, whereas E211Q strongly impairs motility/export. Deleting residues 401–410 retains hydrolysis but uncouples it from productive gate activation. | Site-directed mutagenesis, motility assays, secretion assays, and biochemical ATPase measurements | More than 80% of E211D cells remained motile, swimming at about 50% of wild-type speed; Δ401–410 retained approximately 40% of wild-type ATPase activity |
| Functional interpretation | ATP turnover is not stoichiometrically coupled to each exported polypeptide. Infrequent FliI hydrolysis can switch the gate into a processive, proton-motive-force-driven state, while FliH₂–FliI cycling supports sequential substrate transfer. | Direct Salmonella export experiments and expert synthesis | ATP binding in E211Q was associated with an approximately 80° modeled FliJ rotational step, but hydrolysis was required to reset the cycle |
| 2024 update | FliH/FliI help the FlhA cytoplasmic ring remodel during the rod/hook-to-filament substrate-specificity switch and correct substrate-recognition errors. Separate work showed that a newly identified C-terminal signal can target early substrates without the ATPase or C-ring, demonstrating that FliI facilitates—but is not universally required for—initial targeting. | 2024 Salmonella mutational, secretion, motility, and substrate-targeting studies | Switching normally occurs when the hook reaches approximately 55 nm; early substrates include FlgB/C/D/E/F/G, FliE/J/K, whereas late substrates include FlgK/L, FliC, and FliD |
Table: A concise evidence matrix distinguishing the experimentally supported enzymatic, structural, spatial, and mechanistic properties of Salmonella Typhimurium FliI from inference. Quantitative mutant and 2024 substrate-ordering results highlight that FliI activates and optimizes export rather than continuously powering every translocation event.
FliI is an ATP phosphohydrolase. Its experimentally supported nucleotide substrate is ATP, yielding ADP and inorganic phosphate. The six-subunit FliI ring contains six catalytic sites located at interfaces between neighboring FliI subunits. ATP-dependent oligomerization and intersubunit catalysis are characteristic of its ATPase α/β-chain-related fold. (minamino2022insightintodistinct pages 1-2)
The evidence assembled here does not establish physiologically important hydrolysis of GTP or other nucleoside triphosphates. Thus, ATP should be recorded as the demonstrated substrate; claims of broad NTPase specificity would be insufficiently supported.
Glu211 is a critical catalytic residue examined through E211D and E211Q substitutions. E211D supports only very infrequent ATP hydrolysis yet retains substantial export, whereas E211Q permits an ATP-binding-associated conformational event but strongly compromises productive cycling. In the E211Q context, ATP binding was associated with an approximately 80° modeled rotation of the central FliJ stalk; hydrolysis was interpreted as necessary to reset the complex for another cycle. (minamino2014thebacterialflagellar pages 5-7, minamino2014thebacterialflagellar pages 1-2)
Residues 401–410 are important for coupling ATPase activity to export-gate activation rather than for catalysis alone. Deleting this segment retained approximately 40% of wild-type ATPase activity, and the mutant still interacted with FlhA, FlhB, FliJ, and a FliT–FliD chaperone–substrate complex, yet flagellar assembly and motility were severely impaired. This uncoupling result is strong evidence that bulk ATP hydrolysis is not itself sufficient: FliI must transmit the resulting conformational change productively to FliJ and the gate. (minamino2014thebacterialflagellar pages 4-5, minamino2014thebacterialflagellar pages 5-7)
FliI is a soluble cytoplasmic/peripheral component, not an integral membrane protein and not part of the extracellular flagellum. It occupies two interconverting pools:
Fluorescence imaging of FliI–YFP showed that approximately 90% of basal-body spots exchanged with the freely diffusing pool, demonstrating highly dynamic rather than permanently fixed localization. FliH connects the ATPase to FlhA and to FliN in the cytoplasmic C-ring. (minamino2014thebacterialflagellar pages 4-5, minamino2014thebacterialflagellar pages 1-2)
The assembled ATPase has the stoichiometry FliH₁₂–FliI₆–FliJ₁. Six FliH dimers bind the N-terminal regions of six FliI subunits; FliJ occupies the central pore as a stalk. The related structural estimate describes a ring approximately 10 nm across with a 2.5–3.0 nm central cavity, although those dimensions should be regarded as structure-level support rather than a direct localization measurement in intact LT2 cells. (minamino2022insightintodistinct pages 1-2, narayan2019computationalstudiesto pages 1-4)
The fT3SS constructs the axial flagellar structure by exporting unfolded subunits from the cytoplasm across the inner membrane. The proteins then move through the narrow central channel of the growing flagellum and polymerize at its distal end. The membrane export complex includes FlhA, FlhB, FliP, FliQ, and FliR; reported stoichiometries include a FlhA nonamer and a FliP₅–FliQ₄–FliR₁ channel assembly. (minamino2022insightintodistinct pages 1-2)
FliI contributes at least three related functions:
Once activated, the Salmonella gate functions primarily as an H⁺/protein antiporter, coupling inward proton flow to outward polypeptide export. Thus, FliI hydrolyzes ATP, but it does not itself transport ATP-derived phosphate energy across the membrane or constitute the translocation pore. (minamino2022insightintodistinct pages 1-2)
The strongest experimental interpretation is no. E211D mutant cells, despite extremely infrequent hydrolysis, retained near-wild-type secretion for several substrates; more than 80% were motile, although swimming speed was approximately 50% of wild type. Protein export can therefore continue processively after rare ATPase events. ATP turnover rate is not directly proportional to export rate. (minamino2014thebacterialflagellar pages 1-2)
Genetic experiments further showed that increased PMF and increased substrate abundance can partially bypass loss of the FliHIJ complex. FliHIJ deletion nevertheless makes hook–basal-body completion and substrate-specificity switching inefficient and rare. These results support an expert model in which FliI normally accelerates difficult steps—including substrate delivery, chaperone release/unfolding, gate activation, and fidelity—but the ion-powered gate can perform residual export under compensating conditions. (erhardt2014atpaseindependenttypeiiiprotein pages 9-10)
Claims that FliI is always the direct substrate-unfolding motor should consequently be qualified. Unfolding/chaperone-release assistance is supported by genetic and pathway evidence, but PMF-dependent gate activity can compensate, and some initial targeting events occur independently of the ATPase.
A positively charged FliI surface cluster—including Arg26, Arg27, Arg33, Arg76, and Arg93—contributes to controlled hexamer formation and efficient substrate entry. Arg33 and Arg76 are especially implicated in ring assembly. FlhB gain-of-function mutations that elevate substrate-entry probability restore export to compromised FliI/FliH backgrounds, linking FliI oligomerization to gate access rather than merely ATP turnover. (kinoshita2021apositivecharge pages 1-2)
The direct biological process is flagellar axial-structure assembly, which enables swimming motility. Early exported substrates build the rod, hook, and hook cap; after the hook reaches approximately 55 nm, FliK–FlhB signaling remodels the export apparatus so that late substrates build the hook–filament junction, filament, and cap. (bryant2024identificationofa pages 1-3, kinoshita2024flihandflii pages 1-2)
Loss or severe impairment of FliI causes defective secretion of flagellar building blocks, inefficient hook–basal-body completion, reduced or absent filament formation, and impaired motility. These are consequences of failure to activate and supply the export gate, rather than evidence that FliI is a mature structural constituent of the flagellum. Residual export in ATPase-deficient backgrounds explains why some suppressor conditions can produce wild-type-length or even longer filaments despite inefficient initiation or switching. (erhardt2014atpaseindependenttypeiiiprotein pages 9-10, minamino2014thebacterialflagellar pages 1-2)
The December 2023 EcoSal Plus review describes the Salmonella flagellum as a basal-body motor, universal-joint hook, and helical propeller assembled by a type III secretion apparatus at the flagellar base. It integrates recent cryo-EM structures with classical genetics and places the export ATPase within a multi-stage cellular nanomachine rather than treating it as an isolated ATP-consuming enzyme. DOI: https://doi.org/10.1128/ecosalplus.esp-0011-2023; published December 2023. (minamino2023structureassemblyand pages 12-13)
Kinoshita and colleagues showed that FliH/FliI help the FlhA cytoplasmic nonamer undergo productive remodeling during the rod/hook-to-filament specificity switch. Experiments on the conserved FlhA GYXLI region led the authors to conclude that the ATPase complex both promotes efficient switching and helps correct substrate-recognition errors. This extends FliI’s annotation from “ATPase/gate activator” to a contributor to the fidelity and temporal ordering of assembly. DOI: https://doi.org/10.1038/s42003-024-06081-0; published March 2024. (kinoshita2024flihandflii pages 1-2)
Bryant and Fraser identified a C-terminal targeting signal in early flagellar subunits. This signal could direct substrates to the core export machinery without the flagellar ATPase or cytoplasmic C-ring. The result does not make FliI dispensable under normal conditions; rather, it shows that substrate targeting is multistep and that FliI-mediated delivery is one facilitating route among several sequential recognition mechanisms. DOI: https://doi.org/10.1128/mbio.03067-23; published 20 February 2024. (bryant2024identificationofa pages 1-3)
Together, the 2024 studies refine the current model: FliI is crucial for efficient activation, delivery, and quality control, but neither every initial docking event nor each translocation step requires a contemporaneous ATP-hydrolysis event.
FliI and the fT3SS are widely used as experimental systems for:
The 2024 export-signal experiments used Salmonella motility halos, secretion immunoblots, and defined FlgE variants to map sequential substrate-recognition events, illustrating a current practical implementation of the system in mechanistic protein-export research. (bryant2024identificationofa pages 1-3)
T3SS ATPases are attractive anti-virulence targets because related secretion systems support bacterial motility or delivery of virulence factors. FliI offers a tractable ATPase model for inhibitor discovery and for understanding conserved T3SS ATPases. However, FliI P26465 is not a validated clinical drug target, and the ability of elevated PMF or altered substrate abundance to bypass ATPase loss warns that ATPase inhibition alone may not fully block secretion in every physiological state. Moreover, compounds directed at homologous injectisome ATPases cannot automatically be assumed to inhibit Salmonella FliI selectively. (erhardt2014atpaseindependenttypeiiiprotein pages 9-10)
No approved therapy, diagnostic, industrial process, or clinical trial specifically implementing inhibition or engineering of P26465 was identified in the retrieved evidence.
High confidence: identity as Salmonella flagellar FliI; ATP hydrolysis; soluble cytoplasmic/basal-body localization; FliI₆ ring; FliH₁₂–FliI₆–FliJ₁ architecture; role in efficient flagellar export and motility; interaction with FliH/FliJ/FlhA/FlhB; and separation between ATP turnover and continuous PMF-driven translocation. These points have direct Salmonella genetic, biochemical, imaging, and structural support. (kinoshita2021apositivecharge pages 1-2, minamino2022insightintodistinct pages 1-2, minamino2014thebacterialflagellar pages 1-2)
Moderate confidence/mechanistic model: ATP-driven FliJ rotation and detailed allocation of unfolding, chaperone release, gate opening, and reset steps. The combined evidence strongly supports conformational coupling, but some details remain model-dependent and different experiments emphasize either ATPase-assisted unfolding or gate activation. (erhardt2014atpaseindependenttypeiiiprotein pages 9-10, minamino2014thebacterialflagellar pages 5-7)
Not established: physiologically important alternative nucleotide substrates; a one-ATP-per-exported-subunit stoichiometry; direct membrane transport by FliI; permanent incorporation into the mature extracellular flagellum; or a clinically validated FliI inhibitor.
FliI (P26465/STM1972) is a cytoplasmic, basal-body-associated, homohexameric ATPase of the Salmonella Typhimurium flagellar type III secretion system. It hydrolyzes ATP at six intersubunit catalytic sites and works with FliH and FliJ to deliver flagellar substrates, activate and reset the FlhA/FlhB-associated export gate, and preserve the efficiency and correct order of flagellar assembly. The membrane gate—not FliI itself—conducts substrates and normally uses proton motive force for processive export.
References
(minamino2022insightintodistinct pages 1-2): Tohru Minamino, Miki Kinoshita, and Keiichi Namba. Insight into distinct functional roles of the flagellar atpase complex for flagellar assembly in salmonella. Frontiers in Microbiology, May 2022. URL: https://doi.org/10.3389/fmicb.2022.864178, doi:10.3389/fmicb.2022.864178. This article has 41 citations and is from a peer-reviewed journal.
(minamino2014thebacterialflagellar pages 1-2): Tohru Minamino, Yusuke V. Morimoto, Miki Kinoshita, Phillip D. Aldridge, and Keiichi Namba. The bacterial flagellar protein export apparatus processively transports flagellar proteins even with extremely infrequent atp hydrolysis. Scientific Reports, Dec 2014. URL: https://doi.org/10.1038/srep07579, doi:10.1038/srep07579. This article has 106 citations and is from a peer-reviewed journal.
(kinoshita2021apositivecharge pages 1-2): Miki Kinoshita, Keiichi Namba, and Tohru Minamino. A positive charge region of salmonella flii is required for atpase formation and efficient flagellar protein export. Communications Biology, Apr 2021. URL: https://doi.org/10.1038/s42003-021-01980-y, doi:10.1038/s42003-021-01980-y. This article has 24 citations and is from a peer-reviewed journal.
(OpenTargets Search: -fliI): Open Targets Query (-fliI, 8 results). Buniello, A. et al. (2025). Open Targets Platform: facilitating therapeutic hypotheses building in drug discovery. Nucleic Acids Research.
(minamino2014thebacterialflagellar pages 5-7): Tohru Minamino, Yusuke V. Morimoto, Miki Kinoshita, Phillip D. Aldridge, and Keiichi Namba. The bacterial flagellar protein export apparatus processively transports flagellar proteins even with extremely infrequent atp hydrolysis. Scientific Reports, Dec 2014. URL: https://doi.org/10.1038/srep07579, doi:10.1038/srep07579. This article has 106 citations and is from a peer-reviewed journal.
(minamino2014thebacterialflagellar pages 4-5): Tohru Minamino, Yusuke V. Morimoto, Miki Kinoshita, Phillip D. Aldridge, and Keiichi Namba. The bacterial flagellar protein export apparatus processively transports flagellar proteins even with extremely infrequent atp hydrolysis. Scientific Reports, Dec 2014. URL: https://doi.org/10.1038/srep07579, doi:10.1038/srep07579. This article has 106 citations and is from a peer-reviewed journal.
(narayan2019computationalstudiesto pages 1-4): P Narayan and G NagendraH. Computational studies to decipher the plausible. Unknown journal, 2019.
(kinoshita2024flihandflii pages 1-2): Miki Kinoshita, Tohru Minamino, Takayuki Uchihashi, and Keiichi Namba. Flih and flii help flha bring strict order to flagellar protein export in salmonella. Communications Biology, Mar 2024. URL: https://doi.org/10.1038/s42003-024-06081-0, doi:10.1038/s42003-024-06081-0. This article has 18 citations and is from a peer-reviewed journal.
(erhardt2014atpaseindependenttypeiiiprotein pages 9-10): Marc Erhardt, Max E. Mertens, Florian D. Fabiani, and Kelly T. Hughes. Atpase-independent type-iii protein secretion in salmonella enterica. PLoS Genetics, 10:e1004800, Nov 2014. URL: https://doi.org/10.1371/journal.pgen.1004800, doi:10.1371/journal.pgen.1004800. This article has 106 citations and is from a domain leading peer-reviewed journal.
(bryant2024identificationofa pages 1-3): Owain J. Bryant and Gillian M. Fraser. Identification of a new export signal that targets early subunits to the flagellar type iii secretion export machinery. mBio, Mar 2024. URL: https://doi.org/10.1128/mbio.03067-23, doi:10.1128/mbio.03067-23. This article has 1 citations and is from a domain leading peer-reviewed journal.
(minamino2023structureassemblyand pages 12-13): Tohru Minamino and Miki Kinoshita. Structure, assembly, and function of flagella responsible for bacterial locomotion. EcoSal Plus, Dec 2023. URL: https://doi.org/10.1128/ecosalplus.esp-0011-2023, doi:10.1128/ecosalplus.esp-0011-2023. This article has 69 citations.