TtgA is the N-terminally lipidated periplasmic adaptor or membrane fusion protein subunit of the tripartite TtgABC RND efflux system in Pseudomonas putida KT2440. It is anchored to the inner membrane and helps couple the inner-membrane transporter TtgB to the outer-membrane exit duct TtgC, thereby supporting export of toxic hydrophobic compounds. Evidence from KT2440 and closely related Pseudomonas systems links TtgABC to tolerance against antibiotics, organic solvents, and monoterpenoid toxicants, although the exact substrate range of the KT2440 pump remains incompletely resolved.
Definition: Binding-based adaptor activity by a periplasmic membrane-fusion protein subunit that bridges an inner-membrane RND transporter and an outer-membrane exit duct, stabilizing a tripartite efflux complex that exports toxic compounds across the cell envelope.
Justification: Current GO annotations tend to over-annotate membrane-fusion/adaptor subunits like TtgA as transmembrane transporter activity even though these proteins do not themselves form the transport pore. A dedicated adaptor term would better represent the molecular role of RND pump MFP subunits.
Parent term: transmembrane transporter binding
Supporting Evidence:
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
|
GO:0005886
plasma membrane
|
IEA
GO_REF:0000120 |
ACCEPT |
Summary: This localization is appropriate for TtgA. UniProt identifies TtgA as a lipid-anchored inner-membrane protein, and the membrane-fusion/adaptor subunit of the TtgABC pump is expected to remain associated with the plasma membrane while projecting into the periplasm.
Reason: TtgA is not a soluble periplasmic protein; it is a lipoprotein tethered to the inner membrane. GO:0005886 is therefore an informative and biologically correct cellular component term for this gene product.
Supporting Evidence:
file:PSEPK/ttgA/ttgA-uniprot.txt
SUBCELLULAR LOCATION: Cell inner membrane {ECO:0000305}; Lipid-anchor {ECO:0000255|PROSITE-ProRule:PRU00303}
|
|
GO:0016020
membrane
|
IEA
GO_REF:0000002 |
REMOVE |
Summary: This annotation is too broad. TtgA is membrane-associated, but GO:0005886 already captures the specific and relevant location.
Reason: Retaining the generic membrane term adds little value once the more specific plasma membrane annotation is accepted. The broad InterPro-derived term obscures the fact that TtgA is specifically anchored to the bacterial inner membrane.
|
|
GO:0022857
transmembrane transporter activity
|
IEA
GO_REF:0000002 |
MODIFY |
Summary: This is an over-annotation for TtgA itself. TtgA is the periplasmic adaptor subunit of the TtgABC complex, not the inner-membrane transport pore or the energy-coupling RND transporter. Its role is to stabilize coupling between the transporter and the outer-membrane channel.
Reason: The tripartite pump as a whole enables transmembrane transport, but that activity should not be assigned directly to the non-transmembrane adaptor subunit. A better molecular-function proxy is GO:0044325 transmembrane transporter binding, with complex membership captured separately by GO:1902495.
Proposed replacements:
transmembrane transporter binding
Supporting Evidence:
file:PSEPK/ttgA/ttgA-uniprot.txt
Probable periplasmic linker protein component of the TtgABC
file:PSEPK/ttgA/ttgA-uniprot.txt
Belongs to the membrane fusion protein (MFP) (TC 8.A.1) family.
|
|
GO:0046677
response to antibiotic
|
IEA
GO_REF:0000118 |
KEEP AS NON CORE |
Summary: The annotation is plausible because TtgABC-family pumps contribute to antibiotic tolerance, but it is narrower than the best-supported biological picture. TtgABC systems also handle organic solvents and monoterpenoid toxicants, so antibiotic response is only one facet of a broader xenobiotic-defense role.
Reason: Experimental work in Pseudomonas links TtgABC to antibiotic efflux, but the broader core function is export of multiple toxic hydrophobic compounds. Keeping this term as non-core preserves a supported substrate class without mistaking it for the full functional scope.
Supporting Evidence:
file:PSEPK/ttgA/ttgA-uniprot.txt
GO; GO:0046677; P:response to antibiotic; IEA:TreeGrafter.
|
|
GO:0055085
transmembrane transport
|
IEA
GO_REF:0000120 |
MODIFY |
Summary: The general process term is directionally correct but too broad. The best-supported process is transport of toxic xenobiotic compounds rather than abstract transmembrane transport.
Reason: TtgA functions in a multidrug or solvent-efflux context, so GO:0042908 xenobiotic transport better captures the biological process than the parent term GO:0055085.
Proposed replacements:
xenobiotic transport
Supporting Evidence:
file:PSEPK/ttgA/ttgA-uniprot.txt
GO; GO:0042908; P:xenobiotic transport; IEA:UniProtKB-ARBA.
file:PSEPK/ttgA/ttgA-deep-research-falcon.md
encodes the **membrane fusion protein (MFP) / periplasmic adaptor** subunit
|
|
GO:1902495
transmembrane transporter complex
|
TAS
PMID:32296906 Investigation of monoterpenoid resistance mechanisms in Pseu... |
NEW |
Summary: TtgA is described as the lipoprotein/adaptor component of the tripartite TtgABC transporter assembly. The evidence here is a traceable literature statement and UniProt family assignment, not a direct complex-assembly assay.
Reason: Complex membership is a more accurate representation of TtgA than assigning the subunit the full transporter activity. TAS is used instead of IMP because the GS1 monoterpenoid paper states the Ttg pump architecture and TtgA/D/G adaptor role, while the mutant phenotypes do not directly demonstrate complex assembly.
Supporting Evidence:
PMID:32296906
All three efflux pumps consist of three components: an inner membrane protein (TtgB/E/H), which binds the substrates and acts as the extrusion element, an outer membrane protein that reaches into the periplasmic space to form a channel (TtgC/F/I), and a lipoprotein that plays a role in stabilizing the interaction between the other two elements (TtgA/D/G)
file:PSEPK/ttgA/ttgA-uniprot.txt
Probable periplasmic linker protein component of the TtgABC
|
|
GO:0042908
xenobiotic transport
|
IMP
PMID:32296906 Investigation of monoterpenoid resistance mechanisms in Pseu... |
NEW |
Summary: This is the most useful process-level summary of TtgA function. TtgABC-family pumps protect Pseudomonas cells against chemically diverse toxic hydrophobic compounds including antibiotics, solvents, and monoterpenoids.
Reason: The phenotype of ttgA loss and the broader TtgABC literature both support a role in transport-based detoxification of xenobiotic compounds. This is more precise than the parent term transmembrane transport and more general than the substrate-specific antibiotic-response annotation.
Supporting Evidence:
file:PSEPK/ttgA/ttgA-uniprot.txt
GO; GO:0042908; P:xenobiotic transport; IEA:UniProtKB-ARBA.
|
|
GO:0044325
transmembrane transporter binding
|
TAS
PMID:32296906 Investigation of monoterpenoid resistance mechanisms in Pseu... |
NEW |
Summary: GO currently lacks a term tailored to RND efflux-pump adaptor subunits. As a conservative proxy, transmembrane transporter binding captures the traceable literature statement that the TtgA/D/G lipoprotein class stabilizes the interaction between inner-membrane transporter and outer-membrane channel elements.
Reason: TtgA does not itself span the membrane or catalyze transport. TAS is used instead of ISS because the support is a traceable author statement about TtgA/D/G adaptor interactions, not transfer from a characterized ortholog. Until a more specific adaptor term exists, GO:0044325 is the closest informative molecular-function representation.
Supporting Evidence:
PMID:32296906
a lipoprotein that plays a role in stabilizing the interaction between the other two elements (TtgA/D/G)
file:PSEPK/ttgA/ttgA-uniprot.txt
Probable periplasmic linker protein component of the TtgABC
file:PSEPK/ttgA/ttgA-uniprot.txt
Belongs to the membrane fusion protein (MFP) (TC 8.A.1) family.
|
Q: What is the native substrate spectrum of the KT2440 TtgABC pump under physiological conditions: antibiotics, aromatic solvents, monoterpenoids, or a broader xenobiotic set?
Suggested experts: Ana Segura, Juan L Ramos, Markus Buchhaupt
Q: Does TtgA merely stabilize the TtgB-TtgC assembly, or does it also influence substrate selectivity and efflux efficiency?
Suggested experts: Ana Segura, Juan L Ramos, Markus Buchhaupt
Experiment: Construct clean ttgA and ttgABC deletion mutants in KT2440, complement them in trans, and measure tolerance or MIC phenotypes for chloramphenicol, tetracycline, toluene, 1,8-cineole, verbenone, and geraniol together with a resazurin efflux assay.
Hypothesis: TtgA is required for efficient efflux of multiple toxic hydrophobic compounds in KT2440.
Type: targeted gene deletion and phenotype profiling
Experiment: Mutate the lipobox cysteine and predicted interfacial residues of TtgA, then test complex formation with TtgB and TtgC by co-immunoprecipitation or crosslinking and compare efflux activity against the wild-type protein.
Hypothesis: The lipidated periplasmic adaptor domain of TtgA is required to couple TtgB and TtgC into a functional tripartite complex.
Type: structure-function analysis of efflux-pump assembly
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 gene ttgA (ordered locus PP_1386, UniProt Q88N30) in Pseudomonas putida KT2440 encodes the membrane fusion protein (MFP) / periplasmic adaptor subunit of the tripartite RND (resistance–nodulation–division) efflux pump TtgABC. TtgA physically couples the inner-membrane RND transporter (TtgB) to the outer-membrane channel (TtgC), enabling export of diverse toxic compounds from the inner membrane/periplasm to the external milieu. In KT2440, genetic and transcriptomic evidence links ttgA/ttgABC to clinically relevant β-lactam resistance, resistance to several other antibiotics, and detoxification of non-antibiotic toxicants (e.g., bipyridyl chelators), with strong quantitative MIC phenotypes upon deletion or overexpression. (puja2020coordinateoverexpressionof pages 1-2, tettmann2014knockoutofextracytoplasmic pages 4-5, puja2020coordinateoverexpressionof pages 2-4)
Gene symbol/protein match. In KT2440, ttgA is PP_1386 and is explicitly annotated as an “RND family efflux transporter MFP subunit” (i.e., a membrane fusion protein/periplasmic adaptor) in transcriptome-linked gene tables. This matches the UniProt description “Probable efflux pump periplasmic linker TtgA; precursor” and the expected MFP role in a tripartite RND efflux pump. (tettmann2014knockoutofextracytoplasmic pages 4-5)
Organism match. The quantitative MIC and RNA-seq evidence explicitly refers to **
Pseudomonas putida KT2440**, the same strain context as the UniProt entry provided. (tettmann2014knockoutofextracytoplasmic pages 4-5, puja2020coordinateoverexpressionof pages 2-4)
Family/domain match (conceptual). Multiple KT2440 sources describe TtgA as the periplasmic adaptor/MFP of a tripartite RND efflux system, consistent with membership in the MFP family (TC 8.A.1) and its multi-domain architecture typical of RND pump periplasmic linkers. (puja2020coordinateoverexpressionof pages 1-2, tettmann2014knockoutofextracytoplasmic pages 1-2, tettmann2014knockoutofextracytoplasmic pages 4-5)
Tripartite RND systems form a continuous conduit across the Gram-negative envelope. In KT2440, TtgABC consists of:
- TtgB: inner-membrane RND transporter (the energy-coupled exporter)
- TtgA: periplasmic membrane fusion protein (MFP) / adaptor
- TtgC: outer-membrane channel (outer membrane factor)
This architecture allows substrates to be exported from the periplasm and/or inner membrane to the extracellular milieu. (puja2020coordinateoverexpressionof pages 1-2, tettmann2014knockoutofextracytoplasmic pages 1-2, tettmann2014knockoutofextracytoplasmic pages 4-5)
In the TtgABC complex, TtgA is not the energizing transporter; rather, it is the periplasmic linker/adaptor that mediates assembly and functional coupling between the inner-membrane transporter and the outer-membrane channel. This adaptor role is central to creating the trans-envelope efflux route characteristic of RND systems. (puja2020coordinateoverexpressionof pages 1-2, tettmann2014knockoutofextracytoplasmic pages 4-5)
Primary function: TtgA functions as the MFP/adaptor subunit of the TtgABC RND efflux system, enabling efflux of diverse toxic compounds (antibiotics and non-antibiotics). (puja2020coordinateoverexpressionof pages 1-2, tettmann2014knockoutofextracytoplasmic pages 4-5)
Mechanistic role: TtgABC exports substrates from the periplasm or inner membrane to the extracellular environment, and TtgA is a required structural/functional component for transporter–channel coupling in this tripartite assembly. (puja2020coordinateoverexpressionof pages 1-2)
For KT2440, ttgA (PP_1386) is annotated as the MFP subunit of an RND efflux transporter, placing TtgA primarily in the periplasm, associated with the inner-membrane transporter and contacting the outer-membrane channel as part of the assembled complex. (tettmann2014knockoutofextracytoplasmic pages 4-5)
Because TtgA is an essential subunit of TtgABC, the most defensible statement about “TtgA substrates” is: TtgA supports export of the TtgABC substrate set.
Antibiotics and xenobiotics (KT2440 evidence): TtgABC contributes to resistance against multiple antibiotic classes, with particularly strong effects for several antipseudomonal β-lactams and measurable effects for fluoroquinolones; aminoglycoside effects are limited or absent in certain genetic backgrounds. (puja2020coordinateoverexpressionof pages 2-4, puja2020coordinateoverexpressionof pages 4-5)
Non-antibiotic toxicants (KT2440 evidence): TtgABC is required for detoxification/resistance to 2,2′-bipyridyl and related bipyridyl derivatives (including the natural product caerulomycin A). This suggests a plausible native ecological role in resisting chelators produced by other organisms and maintaining metal homeostasis. (henriquez2020resistancetobipyridyls pages 1-2)
Solvents/aromatic hydrocarbons (context from P. putida efflux literature; strain note): Authoritative reviews and primary studies in the closely related solvent-tolerant strain DOT-T1E link TtgABC to extrusion of aromatic solvents such as toluene, isopropylbenzene, p-xylene and to antibiotic efflux; however, these DOT-T1E findings should be treated as contextual support rather than direct KT2440 proof. (duque2001globalandcognate pages 2-3, segura2014toluenetolerancesystems pages 9-12)
Across P. putida TtgABC literature, TtgR is a TetR-family repressor encoded divergently from ttgA and controlling the ttgABC operon. In DOT-T1E, inactivation of ttgR yields strong derepression of the ttgA promoter (~10–20-fold), establishing TtgR as the key local negative regulator. (duque2001globalandcognate pages 2-3, rojas2003invivoand pages 1-2)
In KT2440, knockout of the extracytoplasmic-function sigma factor ECF-10 caused strong upregulation of ttgA (PP_1386) (~5.9-fold) along with ttgB and ttgC, linking envelope stress regulation to increased efflux capacity. (tettmann2014knockoutofextracytoplasmic pages 4-5)
A key insight from KT2440 multidrug-resistance studies is that efflux phenotypes can reflect coordinated expression of multiple RND systems, not just TtgABC alone. For example, MDR mutants can show TtgABC overproduction due to regulatory mutations, and combined contributions of distinct pumps shape the full resistance profile. (puja2020coordinateoverexpressionof pages 2-4, puja2020coordinateoverexpressionof pages 4-5)
Tettmann et al. (Applied and Environmental Microbiology; publication date: Aug 2014; URL: https://doi.org/10.1128/AEM.01291-14) provides KT2440 MIC tables that directly connect ttgA to antibiotic susceptibility and show transcriptional upregulation of ttgA/ttgB/ttgC.
Key observations:
- In ΔECF-10, ttgA (PP_1386) is upregulated ~5.90-fold (RNA-seq), with ttgB ~4.17-fold and ttgC ~2.94-fold. (tettmann2014knockoutofextracytoplasmic pages 4-5)
- The ttgA mutant shows reduced MICs relative to wild type for multiple antibiotics (examples): ciprofloxacin 0.03→0.015 µg/mL; meropenem 0.03→0.015; chloramphenicol 16→8; nalidixic acid 4→1; sulfamethoxazole 64→16 µg/mL. (tettmann2014knockoutofextracytoplasmic pages 4-5)
These MIC values and the PP_1386 functional annotation are visible in the extracted tables. (tettmann2014knockoutofextracytoplasmic media f0f78e33, tettmann2014knockoutofextracytoplasmic media 8a52eb29)
Puja et al. (Environmental Microbiology; publication date: Sep 2020; URL: https://doi.org/10.1111/1462-2920.15200) provides large-magnitude MIC shifts in KT2440 upon genetic perturbation of the TtgABC system.
Examples from their KT2440 dataset:
- ΔttgAB drastically sensitizes KT2440 to key β-lactams: ticarcillin 128→4 µg/mL (32-fold), piperacillin 8→0.25 (32-fold), aztreonam 16→0.25 (64-fold), with smaller effects on ciprofloxacin (0.06→0.03, 2-fold). (puja2020coordinateoverexpressionof pages 2-4)
- TtgABC overexpression yields strong resistance increases, e.g. ticarcillin 128→1024 µg/mL and ceftazidime 2→8 µg/mL. (puja2020coordinateoverexpressionof pages 2-4)
- In MDR mutants, ttgA can be overexpressed on the order of ~3.8–5.4-fold relative to KT2440, consistent with efflux-driven multidrug resistance mechanisms. (puja2020coordinateoverexpressionof pages 1-2)
Henríquez et al. (Frontiers in Microbiology; publication date: Aug 2020; URL: https://doi.org/10.3389/fmicb.2020.01974) provides physiological evidence that TtgABC is required for detoxification of 2,2′-bipyridyl and natural bipyridyl derivatives, and that loss of the transporter subunit impacts energy status and siderophore/iron acquisition programs.
Key observations:
- A ttgB deletion renders KT2440 susceptible to 2,2′-bipyridyl and caerulomycin A.
- The mutant shows reduced ATP levels and reduced expression of iron acquisition/pyoverdine-related genes; copper addition restores growth and pyoverdine production. (henriquez2020resistancetobipyridyls pages 1-2)
These results support an ecological/physiological role for TtgABC (and by extension its adaptor TtgA) in resisting environmental chelators and maintaining metal homeostasis. (henriquez2020resistancetobipyridyls pages 1-2)
While not focused on TtgA directly, recent KT2440 work highlights that tripartite efflux systems form interacting networks with overlapping substrate spaces; this influences how single-pump knockouts manifest phenotypes and how cells partition roles among pumps for physiology and resistance. (puja2020coordinateoverexpressionof pages 1-2)
A 2023 KT2440 study on another RND pump (ParXY) explicitly frames tripartite systems as transporting not only antibiotics but also toxic metabolites and secreted products, reinforcing the broader physiological framing relevant to interpreting TtgABC/TtgA functions. (puja2020coordinateoverexpressionof pages 1-2)
In the context of engineering P. putida for production of industrial aromatics, recent work emphasizes induction and use of efflux pumps to manage toxicity of produced compounds and intermediates, with evidence that distinct RND pumps can dominate depending on the compound and strain context. This supports real-world relevance of efflux components like TtgA in strain robustness strategies (noting that some 2023 work emphasizes other pumps such as TtgGHI in solvent-tolerant backgrounds). (franco2025engineeringpseudomonasputida pages 57-60)
Coverage limitation (2024): In the retrieved/accessible corpus here, I did not identify a 2024 primary research article specifically dedicated to KT2440 ttgA (PP_1386); therefore, the newest direct quantitative KT2440 ttgA evidence available in this report is primarily 2014–2020, with 2023 sources contributing contextual advances in efflux network and application framing. (franco2025engineeringpseudomonasputida pages 57-60, tettmann2014knockoutofextracytoplasmic pages 4-5, puja2020coordinateoverexpressionof pages 2-4)
KT2440 and clinical/environmental P. putida isolates can display significant β-lactam and fluoroquinolone resistance phenotypes driven by efflux. The magnitude of MIC shifts upon TtgABC perturbation (tens-fold for some β-lactams) indicates that TtgABC is a major determinant shaping susceptibility profiles, relevant to clinical surveillance and to assessing biosafety of engineered P. putida strains. (puja2020coordinateoverexpressionof pages 2-4)
Efflux pumps are frequently exploited (or avoided) in industrial biotechnology for tolerance engineering. Evidence that TtgABC exports diverse toxic compounds and that its expression is regulated by envelope stress responses in KT2440 supports its relevance as a tunable module for robustness—balanced against potential tradeoffs (e.g., energetic costs or altered physiology) implied by stress-regulatory coupling. (tettmann2014knockoutofextracytoplasmic pages 4-5)
A synthesis from reviews and foundational primary studies in P. putida efflux biology frames TtgABC as an RND efflux pump involved in solvent and antibiotic resistance, with TtgR as a key repressor that responds to certain antibiotics/plant metabolites and can strongly derepress the operon when inactivated. Reviews also emphasize that high solvent tolerance generally involves multiple mechanisms/pumps, and that increasing TtgABC expression does not necessarily translate into higher solvent tolerance in all contexts—highlighting the importance of system-level interpretation and strain-specific contributions. (segura2014toluenetolerancesystems pages 9-12)
The following table compiles the most directly supported, KT2440-centered functional annotation and quantitative phenotypes for ttgA/PP_1386 and its cognate pump components.
| Component | Gene / locus | Protein role | Localization | Known / implicated substrates | Regulation | Key quantitative phenotypes | Key references |
|---|---|---|---|---|---|---|---|
| TtgA | ttgA / PP_1386 / UniProt Q88N30 | Periplasmic membrane fusion protein (MFP) / adaptor subunit of the tripartite RND pump TtgABC; links inner-membrane transporter to outer-membrane channel (puja2020coordinateoverexpressionof pages 1-2, tettmann2014knockoutofextracytoplasmic pages 1-2, tettmann2014knockoutofextracytoplasmic pages 4-5) | Periplasmic adaptor associated with the inner membrane–outer membrane efflux assembly; precursor protein per UniProt and MFP annotation in KT2440 (puja2020coordinateoverexpressionof pages 1-2, tettmann2014knockoutofextracytoplasmic pages 4-5) | As part of TtgABC, implicated in export of β-lactams, fluoroquinolones, chloramphenicol, tetracycline, erythromycin, nalidixic acid; aromatic solvents/hydrophobics such as toluene, isopropylbenzene, p-xylene, trichlorobenzene; also flavonoids, bipyridyls/caerulomycin A, bile salts/deoxycholate, and reported nonnative substrates such as styrene/1-naphthol/butanol in broader TtgABC literature (KT2440 unless noted; some solvent data from DOT-T1E) (puja2020coordinateoverexpressionof pages 1-2, duque2001globalandcognate pages 2-3, rojas2003invivoand pages 1-2, puja2020coordinateoverexpressionof pages 2-4, henriquez2020resistancetobipyridyls pages 1-2) | Local: TtgR (TetR-family repressor) divergently transcribed from ttgA; loss of TtgR derepresses ttgABC. Global/other: ECF-10 negatively affects ttgABC expression in KT2440; Lrp-like global regulator affects ttgABC via ttgR in DOT-T1E; PpeRS positively controls TtgABC in KT2440 MDR backgrounds (duque2001globalandcognate pages 2-3, duque2001globalandcognate pages 1-2, rojas2003invivoand pages 1-2, tettmann2014knockoutofextracytoplasmic pages 4-5, puja2020coordinateoverexpressionof pages 2-4) | KT2440: ttgA (PP1386) upregulated ~5.90-fold in ΔECF-10; ttgA mutant MICs drop vs WT for CIP 0.03→0.015, MER 0.03→0.015, CM 16→8, NA 4→1, SX 64→16 µg/mL; TOB 0.125→0.06, IMP unchanged 0.125. Overexpression context: ttgA overexpressed ~3.8–5.4-fold in TtgABC-overproducing mutants relative to KT2440. DOT-T1E note: ttgA promoter rises ~10–20-fold in ttgR mutants; ttgA mRNA about 1.5-fold lower with toluene while ttgR mRNA rises 4–8-fold (duque2001globalandcognate pages 2-3, duque2001globalandcognate pages 1-2, rojas2003invivoand pages 1-2, tettmann2014knockoutofextracytoplasmic pages 4-5, puja2020coordinateoverexpressionof pages 2-4) | Tettmann 2014, https://doi.org/10.1128/AEM.01291-14; Puja 2020, https://doi.org/10.1111/1462-2920.15200; Duque 2001 (DOT-T1E), https://doi.org/10.1046/j.1365-2958.2001.02310.x; Rojas 2003 (DOT-T1E), https://doi.org/10.1128/JB.185.16.4755-4763.2003; Henríquez 2020, https://doi.org/10.3389/fmicb.2020.01974 |
| TtgB | ttgB / PP_1385 | Inner-membrane RND transporter / HAE1 family exporter; energy-coupled pump component that interacts with TtgA and TtgC to move substrates from periplasm or inner membrane to outside (puja2020coordinateoverexpressionof pages 1-2, tettmann2014knockoutofextracytoplasmic pages 4-5) | Inner membrane (canonical RND transporter position) (puja2020coordinateoverexpressionof pages 1-2, tettmann2014knockoutofextracytoplasmic pages 1-2, tettmann2014knockoutofextracytoplasmic pages 4-5) | Same TtgABC substrate spectrum as above; direct KT2440 evidence includes resistance to many β-lactams and fluoroquinolones, bipyridyl chelators and related natural products, bile salts/deoxycholate; broader literature also implicates aromatic solvents and hydrophobic toxicants (some solvent data from DOT-T1E) (puja2020coordinateoverexpressionof pages 1-2, duque2001globalandcognate pages 2-3, puja2020coordinateoverexpressionof pages 2-4, henriquez2020resistancetobipyridyls pages 1-2) | Controlled with ttgA/ttgC in the ttgABC operon; repressed by TtgR; upregulated in ΔECF-10 and MDR regulatory backgrounds such as altered PpeRS signaling (duque2001globalandcognate pages 2-3, tettmann2014knockoutofextracytoplasmic pages 4-5, puja2020coordinateoverexpressionof pages 2-4) | KT2440 ΔttgAB: MICs fall strongly: ticarcillin 128→4 (32-fold), piperacillin 8→0.25 (32-fold), aztreonam 16→0.25 (64-fold), ceftazidime 2→0.5 (4-fold), cefepime 1→0.5 (2-fold), meropenem 0.25→0.06 (~4-fold), ciprofloxacin 0.06→0.03 (2-fold), gentamicin unchanged 0.25→0.25. KT2440 ttgB mutant: susceptible to 2,2'-bipyridyl and caerulomycin A; reduced ATP and impaired pyoverdine/iron-acquisition responses, rescued by copper. ΔECF-10: ttgB up ~4.17-fold (tettmann2014knockoutofextracytoplasmic pages 4-5, puja2020coordinateoverexpressionof pages 2-4, henriquez2020resistancetobipyridyls pages 1-2) | Puja 2020, https://doi.org/10.1111/1462-2920.15200; Henríquez 2020, https://doi.org/10.3389/fmicb.2020.01974; Tettmann 2014, https://doi.org/10.1128/AEM.01291-14 |
| TtgC | ttgC / PP_1384 | Outer-membrane factor / NodT-like channel completing the tripartite conduit (puja2020coordinateoverexpressionof pages 1-2, tettmann2014knockoutofextracytoplasmic pages 4-5) | Outer membrane (puja2020coordinateoverexpressionof pages 1-2, tettmann2014knockoutofextracytoplasmic pages 1-2, tettmann2014knockoutofextracytoplasmic pages 4-5) | Same TtgABC-associated substrates; required for full antibiotic/dye resistance profile of the TtgABC system in KT2440; broader solvent role mainly inferred from TtgABC studies including DOT-T1E (puja2020coordinateoverexpressionof pages 1-2, puja2020coordinateoverexpressionof pages 4-5) | Co-regulated in ttgABC operon with TtgA/B; subject to TtgR repression and ECF-10/PpeRS-linked control in KT2440-related studies (tettmann2014knockoutofextracytoplasmic pages 4-5, puja2020coordinateoverexpressionof pages 2-4, puja2020coordinateoverexpressionof pages 4-5) | KT2440: ttgC up ~2.94-fold in ΔECF-10. In-frame ΔttgC phenocopies strong susceptibility effects seen with loss of TtgABC/associated channel function, supporting dependence on this outer-membrane channel for resistance to multiple β-lactams, fluoroquinolones, chloramphenicol, tetracycline, novobiocin, and dyes (quantified in ΔttgAB datasets and comparative mutant analyses) (tettmann2014knockoutofextracytoplasmic pages 4-5, puja2020coordinateoverexpressionof pages 4-5) | Tettmann 2014, https://doi.org/10.1128/AEM.01291-14; Puja 2020, https://doi.org/10.1111/1462-2920.15200 |
| TtgABC system note | Operon: PP_1386-PP_1384 | Chromosomally encoded tripartite RND efflux system in Pseudomonas putida KT2440; mechanistically analogous to MexAB-OprM-like assemblies, exporting substrates from inner membrane/periplasm directly to extracellular milieu (puja2020coordinateoverexpressionof pages 1-2, tettmann2014knockoutofextracytoplasmic pages 4-5) | Spans inner membrane, periplasm, and outer membrane (puja2020coordinateoverexpressionof pages 1-2, tettmann2014knockoutofextracytoplasmic pages 1-2) | Native roles include multidrug resistance and detoxification of hydrophobic chemicals; likely contributes to ecological defense against plant metabolites/flavonoids and microbial chelators, and to solvent stress adaptation. In KT2440, evidence is strongest for antibiotics and bipyridyl-family toxicants; in DOT-T1E, solvent/aromatic hydrocarbon export is especially well established (puja2020coordinateoverexpressionof pages 1-2, duque2001globalandcognate pages 2-3, segura2014toluenetolerancesystems pages 9-12, henriquez2020resistancetobipyridyls pages 1-2) | TtgR is the principal local repressor; TtgR recognizes several antibiotics and plant secondary metabolites. Overexpression of TtgABC increases antibiotic resistance, but in DOT-T1E does not by itself measurably increase toluene tolerance, implying other systems such as TtgGHI are more important for extreme solvent tolerance (segura2014toluenetolerancesystems pages 9-12, puja2020coordinateoverexpressionof pages 2-4) | KT2440 overexpression: engineered/selected TtgABC overproducers show major MIC increases, e.g. ticarcillin 128→1024, piperacillin 8→32, aztreonam 16→64, ceftazidime 2→8 µg/mL. KT2440 deletion: large sensitization especially to β-lactams and some quinolones, but not aminoglycosides. DOT-T1E note: TtgABC contributes to intrinsic solvent tolerance but appears accessory relative to TtgGHI for high toluene tolerance (segura2014toluenetolerancesystems pages 9-12, puja2020coordinateoverexpressionof pages 2-4, puja2020coordinateoverexpressionof pages 7-8) | Puja 2020, https://doi.org/10.1111/1462-2920.15200; Segura 2014 (review; DOT-T1E emphasis), https://doi.org/10.1007/978-4-431-54520-0_11; Duque 2001 (DOT-T1E), https://doi.org/10.1046/j.1365-2958.2001.02310.x |
Table: This table summarizes the experimentally supported functional annotation of Pseudomonas putida KT2440 ttgA (PP_1386; UniProt Q88N30) and its partner subunits TtgB/TtgC within the TtgABC RND efflux pump. It highlights subunit roles, localization, substrates, regulation, and quantitative phenotypes while clearly flagging evidence derived from DOT-T1E rather than KT2440.
In Pseudomonas putida KT2440, ttgA (PP_1386; UniProt Q88N30) encodes a periplasmic membrane fusion protein/adaptor required for the activity of the TtgABC tripartite RND efflux pump. The primary biological role of TtgA is structural/functional coupling of TtgB (inner-membrane transporter) to TtgC (outer-membrane channel), enabling efflux of diverse toxic compounds. In KT2440, disruption of ttgA or other TtgABC components yields clear susceptibility phenotypes—especially large MIC reductions for multiple β-lactams—while conditions or mutations that upregulate ttgA/ttgABC increase resistance. The pump additionally contributes to detoxification of non-antibiotic toxicants (e.g., bipyridyl chelators) with downstream effects on cellular energy and iron-acquisition physiology, underscoring a plausible native ecological defense function beyond antibiotics. (tettmann2014knockoutofextracytoplasmic pages 4-5, puja2020coordinateoverexpressionof pages 2-4, henriquez2020resistancetobipyridyls pages 1-2)
References
(puja2020coordinateoverexpressionof pages 1-2): Hélène Puja, Gwendoline Comment, Sophie Chassagne, Patrick Plésiat, and Katy Jeannot. Coordinate overexpression of two
(tettmann2014knockoutofextracytoplasmic pages 4-5): Beatrix Tettmann, Andreas Dötsch, Olivier Armant, Christopher D. Fjell, and Joerg Overhage. Knockout of extracytoplasmic function sigma factor ecf-10 affects stress resistance and biofilm formation in pseudomonas putida kt2440. Aug 2014. URL: https://doi.org/10.1128/aem.01291-14, doi:10.1128/aem.01291-14. This article has 37 citations and is from a peer-reviewed journal.
(puja2020coordinateoverexpressionof pages 2-4): Hélène Puja, Gwendoline Comment, Sophie Chassagne, Patrick Plésiat, and Katy Jeannot. Coordinate overexpression of two
(tettmann2014knockoutofextracytoplasmic pages 1-2): Beatrix Tettmann, Andreas Dötsch, Olivier Armant, Christopher D. Fjell, and Joerg Overhage. Knockout of extracytoplasmic function sigma factor ecf-10 affects stress resistance and biofilm formation in pseudomonas putida kt2440. Aug 2014. URL: https://doi.org/10.1128/aem.01291-14, doi:10.1128/aem.01291-14. This article has 37 citations and is from a peer-reviewed journal.
(puja2020coordinateoverexpressionof pages 4-5): Hélène Puja, Gwendoline Comment, Sophie Chassagne, Patrick Plésiat, and Katy Jeannot. Coordinate overexpression of two
(henriquez2020resistancetobipyridyls pages 1-2): Tania Henríquez, Nicola Victoria Stein, and Heinrich Jung. Resistance to bipyridyls mediated by the ttgabc efflux system in pseudomonas putida kt2440. Frontiers in Microbiology, Aug 2020. URL: https://doi.org/10.3389/fmicb.2020.01974, doi:10.3389/fmicb.2020.01974. This article has 14 citations and is from a peer-reviewed journal.
(duque2001globalandcognate pages 2-3): Estrella Duque, Ana Segura, Gilberto Mosqueda, and Juan L. Ramos. Global and cognate regulators control the expression of the organic solvent efflux pumps ttgabc and ttgdef of pseudomonas putida. Molecular Microbiology, 39:1100-1106, Feb 2001. URL: https://doi.org/10.1046/j.1365-2958.2001.02310.x, doi:10.1046/j.1365-2958.2001.02310.x. This article has 136 citations and is from a domain leading peer-reviewed journal.
(segura2014toluenetolerancesystems pages 9-12): Ana Segura and Juan Luis Ramos. Toluene tolerance systems in pseudomonas. ArXiv, pages 227-248, Sep 2014. URL: https://doi.org/10.1007/978-4-431-54520-0_11, doi:10.1007/978-4-431-54520-0_11. This article has 8 citations.
(rojas2003invivoand pages 1-2): Antonia Rojas, Ana Segura, María Eugenia Guazzaroni, Wilson Terán, Ana Hurtado, María Trinidad Gallegos, and Juan L. Ramos. In vivo and in vitro evidence that ttgv is the specific regulator of the ttgghi multidrug and solvent efflux pump of pseudomonas putida. Journal of Bacteriology, 185:4755-4763, Aug 2003. URL: https://doi.org/10.1128/jb.185.16.4755-4763.2003, doi:10.1128/jb.185.16.4755-4763.2003. This article has 103 citations and is from a peer-reviewed journal.
(tettmann2014knockoutofextracytoplasmic media f0f78e33): Beatrix Tettmann, Andreas Dötsch, Olivier Armant, Christopher D. Fjell, and Joerg Overhage. Knockout of extracytoplasmic function sigma factor ecf-10 affects stress resistance and biofilm formation in pseudomonas putida kt2440. Aug 2014. URL: https://doi.org/10.1128/aem.01291-14, doi:10.1128/aem.01291-14. This article has 37 citations and is from a peer-reviewed journal.
(tettmann2014knockoutofextracytoplasmic media 8a52eb29): Beatrix Tettmann, Andreas Dötsch, Olivier Armant, Christopher D. Fjell, and Joerg Overhage. Knockout of extracytoplasmic function sigma factor ecf-10 affects stress resistance and biofilm formation in pseudomonas putida kt2440. Aug 2014. URL: https://doi.org/10.1128/aem.01291-14, doi:10.1128/aem.01291-14. This article has 37 citations and is from a peer-reviewed journal.
(franco2025engineeringpseudomonasputida pages 57-60): AÁ García Franco. Engineering pseudomonas putida for sustainable production of styrene. Unknown journal, 2025.
(duque2001globalandcognate pages 1-2): Estrella Duque, Ana Segura, Gilberto Mosqueda, and Juan L. Ramos. Global and cognate regulators control the expression of the organic solvent efflux pumps ttgabc and ttgdef of pseudomonas putida. Molecular Microbiology, 39:1100-1106, Feb 2001. URL: https://doi.org/10.1046/j.1365-2958.2001.02310.x, doi:10.1046/j.1365-2958.2001.02310.x. This article has 136 citations and is from a domain leading peer-reviewed journal.
(puja2020coordinateoverexpressionof pages 7-8): Hélène Puja, Gwendoline Comment, Sophie Chassagne, Patrick Plésiat, and Katy Jeannot. Coordinate overexpression of two
id: Q88N30
gene_symbol: ttgA
product_type: PROTEIN
aliases:
- PP_1386
status: DRAFT
taxon:
id: NCBITaxon:160488
label: Pseudomonas putida (strain ATCC 47054 / DSM 6125 / CFBP 8728 / NCIMB 11950
/ KT2440)
description: >-
TtgA is the N-terminally lipidated periplasmic adaptor or membrane fusion protein
subunit of the tripartite TtgABC RND efflux system in Pseudomonas putida KT2440.
It is anchored to the inner membrane and helps couple the inner-membrane transporter
TtgB to the outer-membrane exit duct TtgC, thereby supporting export of toxic
hydrophobic compounds. Evidence from KT2440 and closely related Pseudomonas systems
links TtgABC to tolerance against antibiotics, organic solvents, and monoterpenoid
toxicants, although the exact substrate range of the KT2440 pump remains incompletely
resolved.
references:
- id: GO_REF:0000002
title: Gene Ontology annotation through association of InterPro records with GO
terms
findings:
- statement: InterPro2GO supplies family/domain-based automated annotations
that require curator review against the specific TtgA adaptor protein context.
- id: GO_REF:0000118
title: TreeGrafter-generated GO annotations
findings:
- statement: TreeGrafter transfers GO terms from phylogenetic families; curator
review checks whether the transferred term matches the P. putida TtgA ortholog.
- id: GO_REF:0000120
title: Combined Automated Annotation using Multiple IEA Methods
findings:
- statement: The combined UniProt automated pipeline annotation provides IEA
terms that must be checked for specificity and evidence fit.
- id: PMID:12743835
title: Comparative genomic analysis of solvent extrusion pumps in Pseudomonas
strains exhibiting different degrees of solvent tolerance
findings:
- statement: Solvent tolerance differences across Pseudomonas strains correlate
with the number and type of RND solvent extrusion pumps.
supporting_text: We also studied the number and type of efflux pumps in different
strains in comparison with the P. putida DOT-T1E strain.
- id: PMID:14506010
title: Antibiotic-dependent induction of Pseudomonas putida DOT-T1E TtgABC efflux
pump is mediated by the drug binding repressor TtgR
findings:
- statement: TtgABC is a broad-substrate RND efflux pump whose expression is
induced by hydrophobic antibiotics.
supporting_text: The TtgABC efflux pump, which has a broad substrate specificity,
extrudes antibiotics such as ampicillin, carbenicillin, tetracycline, nalidixic
acid, and chloramphenicol.
- id: PMID:32296906
title: Investigation of monoterpenoid resistance mechanisms in Pseudomonas putida
and their consequences for biotransformations
findings:
- statement: KT2440 carries only the TtgABC member of the Ttg solvent-efflux
systems.
supporting_text: the frequently used KT2440 strain contains only the TtgABC
system
- statement: TtgA is the lipoprotein adaptor subunit of the TtgABC pump.
supporting_text: a lipoprotein that plays a role in stabilizing the interaction
between the other two elements (TtgA/D/G)
- statement: Loss of ttgA lowers efflux activity and compromises tolerance to
several monoterpenoid toxicants.
supporting_text: the deletion in the ttgA gene results in a loss of the TtgABC
efflux system accompanied by a reduced resazurin efflux activity
- id: file:PSEPK/ttgA/ttgA-uniprot.txt
title: UniProt entry Q88N30
findings:
- statement: UniProt identifies TtgA as a probable periplasmic linker protein
component of the TtgABC efflux pump.
- statement: UniProt places TtgA in the membrane fusion protein family and
annotates it as a lipid-anchored inner-membrane protein.
- id: file:PSEPK/ttgA/ttgA-deep-research-falcon.md
title: Falcon deep research report for ttgA in Pseudomonas putida KT2440
findings:
- statement: ttgA encodes the membrane fusion or periplasmic adaptor subunit
of the TtgABC tripartite RND efflux pump in KT2440.
- statement: KT2440 genetic evidence supports a role for TtgABC in efflux-mediated
resistance to antibiotics and other xenobiotic toxicants.
existing_annotations:
- term:
id: GO:0005886
label: plasma membrane
evidence_type: IEA
original_reference_id: GO_REF:0000120
review:
summary: >-
This localization is appropriate for TtgA. UniProt identifies TtgA as a
lipid-anchored inner-membrane protein, and the membrane-fusion/adaptor subunit
of the TtgABC pump is expected to remain associated with the plasma membrane
while projecting into the periplasm.
action: ACCEPT
reason: >-
TtgA is not a soluble periplasmic protein; it is a lipoprotein tethered to
the inner membrane. GO:0005886 is therefore an informative and biologically
correct cellular component term for this gene product.
supported_by:
- reference_id: file:PSEPK/ttgA/ttgA-uniprot.txt
supporting_text: "SUBCELLULAR LOCATION: Cell inner membrane {ECO:0000305}; Lipid-anchor {ECO:0000255|PROSITE-ProRule:PRU00303}"
- term:
id: GO:0016020
label: membrane
evidence_type: IEA
original_reference_id: GO_REF:0000002
review:
summary: >-
This annotation is too broad. TtgA is membrane-associated, but GO:0005886
already captures the specific and relevant location.
action: REMOVE
reason: >-
Retaining the generic membrane term adds little value once the more specific
plasma membrane annotation is accepted. The broad InterPro-derived term obscures
the fact that TtgA is specifically anchored to the bacterial inner membrane.
- term:
id: GO:0022857
label: transmembrane transporter activity
evidence_type: IEA
original_reference_id: GO_REF:0000002
review:
summary: >-
This is an over-annotation for TtgA itself. TtgA is the periplasmic adaptor
subunit of the TtgABC complex, not the inner-membrane transport pore or the
energy-coupling RND transporter. Its role is to stabilize coupling between
the transporter and the outer-membrane channel.
action: MODIFY
reason: >-
The tripartite pump as a whole enables transmembrane transport, but that activity
should not be assigned directly to the non-transmembrane adaptor subunit.
A better molecular-function proxy is GO:0044325 transmembrane transporter
binding, with complex membership captured separately by GO:1902495.
proposed_replacement_terms:
- id: GO:0044325
label: transmembrane transporter binding
supported_by:
- reference_id: file:PSEPK/ttgA/ttgA-uniprot.txt
supporting_text: Probable periplasmic linker protein component of the TtgABC
- reference_id: file:PSEPK/ttgA/ttgA-uniprot.txt
supporting_text: Belongs to the membrane fusion protein (MFP) (TC 8.A.1)
family.
- term:
id: GO:0046677
label: response to antibiotic
evidence_type: IEA
original_reference_id: GO_REF:0000118
review:
summary: >-
The annotation is plausible because TtgABC-family pumps contribute to antibiotic
tolerance, but it is narrower than the best-supported biological picture. TtgABC
systems also handle organic solvents and monoterpenoid toxicants, so antibiotic
response is only one facet of a broader xenobiotic-defense role.
action: KEEP_AS_NON_CORE
reason: >-
Experimental work in Pseudomonas links TtgABC to antibiotic efflux, but the
broader core function is export of multiple toxic hydrophobic compounds. Keeping
this term as non-core preserves a supported substrate class without mistaking
it for the full functional scope.
supported_by:
- reference_id: file:PSEPK/ttgA/ttgA-uniprot.txt
supporting_text: "GO; GO:0046677; P:response to antibiotic; IEA:TreeGrafter."
- term:
id: GO:0055085
label: transmembrane transport
evidence_type: IEA
original_reference_id: GO_REF:0000120
review:
summary: >-
The general process term is directionally correct but too broad. The best-supported
process is transport of toxic xenobiotic compounds rather than abstract transmembrane
transport.
action: MODIFY
reason: >-
TtgA functions in a multidrug or solvent-efflux context, so GO:0042908 xenobiotic
transport better captures the biological process than the parent term GO:0055085.
proposed_replacement_terms:
- id: GO:0042908
label: xenobiotic transport
supported_by:
- reference_id: file:PSEPK/ttgA/ttgA-uniprot.txt
supporting_text: "GO; GO:0042908; P:xenobiotic transport; IEA:UniProtKB-ARBA."
- reference_id: file:PSEPK/ttgA/ttgA-deep-research-falcon.md
supporting_text: encodes the **membrane fusion protein (MFP) / periplasmic adaptor** subunit
- term:
id: GO:1902495
label: transmembrane transporter complex
evidence_type: TAS
original_reference_id: PMID:32296906
review:
summary: >-
TtgA is described as the lipoprotein/adaptor component of the tripartite
TtgABC transporter assembly. The evidence here is a traceable literature
statement and UniProt family assignment, not a direct complex-assembly assay.
action: NEW
reason: >-
Complex membership is a more accurate representation of TtgA than assigning
the subunit the full transporter activity. TAS is used instead of IMP because
the GS1 monoterpenoid paper states the Ttg pump architecture and TtgA/D/G
adaptor role, while the mutant phenotypes do not directly demonstrate complex
assembly.
supported_by:
- reference_id: PMID:32296906
supporting_text: "All three efflux pumps consist of three components: an inner membrane protein (TtgB/E/H), which binds the substrates and acts as the extrusion element, an outer membrane protein that reaches into the periplasmic space to form a channel (TtgC/F/I), and a lipoprotein that plays a role in stabilizing the interaction between the other two elements (TtgA/D/G)"
reference_section_type: INTRODUCTION
- reference_id: file:PSEPK/ttgA/ttgA-uniprot.txt
supporting_text: Probable periplasmic linker protein component of the TtgABC
- term:
id: GO:0042908
label: xenobiotic transport
evidence_type: IMP
original_reference_id: PMID:32296906
review:
summary: >-
This is the most useful process-level summary of TtgA function. TtgABC-family
pumps protect Pseudomonas cells against chemically diverse toxic hydrophobic
compounds including antibiotics, solvents, and monoterpenoids.
action: NEW
reason: >-
The phenotype of ttgA loss and the broader TtgABC literature both support
a role in transport-based detoxification of xenobiotic compounds. This is
more precise than the parent term transmembrane transport and more general
than the substrate-specific antibiotic-response annotation.
supported_by:
- reference_id: file:PSEPK/ttgA/ttgA-uniprot.txt
supporting_text: "GO; GO:0042908; P:xenobiotic transport; IEA:UniProtKB-ARBA."
- term:
id: GO:0044325
label: transmembrane transporter binding
evidence_type: TAS
original_reference_id: PMID:32296906
review:
summary: >-
GO currently lacks a term tailored to RND efflux-pump adaptor subunits. As
a conservative proxy, transmembrane transporter binding captures the traceable
literature statement that the TtgA/D/G lipoprotein class stabilizes the
interaction between inner-membrane transporter and outer-membrane channel
elements.
action: NEW
reason: >-
TtgA does not itself span the membrane or catalyze transport. TAS is used
instead of ISS because the support is a traceable author statement about
TtgA/D/G adaptor interactions, not transfer from a characterized ortholog.
Until a more specific adaptor term exists, GO:0044325 is the closest
informative molecular-function representation.
supported_by:
- reference_id: PMID:32296906
supporting_text: "a lipoprotein that plays a role in stabilizing the interaction between the other two elements (TtgA/D/G)"
reference_section_type: INTRODUCTION
- reference_id: file:PSEPK/ttgA/ttgA-uniprot.txt
supporting_text: Probable periplasmic linker protein component of the TtgABC
- reference_id: file:PSEPK/ttgA/ttgA-uniprot.txt
supporting_text: Belongs to the membrane fusion protein (MFP) (TC 8.A.1)
family.
core_functions:
- molecular_function:
id: GO:0044325
label: transmembrane transporter binding
contributes_to_molecular_function:
id: GO:0022857
label: transmembrane transporter activity
directly_involved_in:
- id: GO:0042908
label: xenobiotic transport
locations:
- id: GO:0005886
label: plasma membrane
in_complex:
id: GO:1902495
label: transmembrane transporter complex
description: >-
TtgA is the inner-membrane-anchored adaptor subunit of the TtgABC tripartite
RND efflux pump. By binding and stabilizing the transporter/channel components,
it contributes to complex-level transmembrane efflux of toxic hydrophobic compounds
across the cell envelope.
supported_by:
- reference_id: file:PSEPK/ttgA/ttgA-uniprot.txt
supporting_text: Probable periplasmic linker protein component of the TtgABC
- reference_id: file:PSEPK/ttgA/ttgA-uniprot.txt
supporting_text: "GO; GO:0042908; P:xenobiotic transport; IEA:UniProtKB-ARBA."
proposed_new_terms:
- proposed_name: RND efflux pump adaptor activity
proposed_definition: >-
Binding-based adaptor activity by a periplasmic membrane-fusion protein subunit
that bridges an inner-membrane RND transporter and an outer-membrane exit duct,
stabilizing a tripartite efflux complex that exports toxic compounds across
the cell envelope.
justification: >-
Current GO annotations tend to over-annotate membrane-fusion/adaptor subunits
like TtgA as transmembrane transporter activity even though these proteins do
not themselves form the transport pore. A dedicated adaptor term would better
represent the molecular role of RND pump MFP subunits.
proposed_parent:
id: GO:0044325
label: transmembrane transporter binding
supported_by:
- reference_id: file:PSEPK/ttgA/ttgA-uniprot.txt
supporting_text: Probable periplasmic linker protein component of the TtgABC
suggested_questions:
- question: >-
What is the native substrate spectrum of the KT2440 TtgABC pump under physiological
conditions: antibiotics, aromatic solvents, monoterpenoids, or a broader xenobiotic
set?
experts:
- Ana Segura
- Juan L Ramos
- Markus Buchhaupt
- question: >-
Does TtgA merely stabilize the TtgB-TtgC assembly, or does it also influence
substrate selectivity and efflux efficiency?
experts:
- Ana Segura
- Juan L Ramos
- Markus Buchhaupt
suggested_experiments:
- hypothesis: TtgA is required for efficient efflux of multiple toxic hydrophobic
compounds in KT2440.
description: >-
Construct clean ttgA and ttgABC deletion mutants in KT2440, complement them
in trans, and measure tolerance or MIC phenotypes for chloramphenicol, tetracycline,
toluene, 1,8-cineole, verbenone, and geraniol together with a resazurin efflux
assay.
experiment_type: targeted gene deletion and phenotype profiling
- hypothesis: The lipidated periplasmic adaptor domain of TtgA is required to couple
TtgB and TtgC into a functional tripartite complex.
description: >-
Mutate the lipobox cysteine and predicted interfacial residues of TtgA, then
test complex formation with TtgB and TtgC by co-immunoprecipitation or crosslinking
and compare efflux activity against the wild-type protein.
experiment_type: structure-function analysis of efflux-pump assembly