SecF is a multi-pass inner membrane protein that forms the SecDF accessory complex together with SecD. As part of the Sec protein translocase, SecDF uses the proton motive force (PMF) to enhance protein translocation across the cytoplasmic membrane after the ATP-dependent function of SecA. SecDF undergoes PMF-driven conformational cycles that pull preproteins through the SecYEG channel into the periplasm, functioning as a membrane-integrated chaperone that works independently of ATP. In P. aeruginosa, the SecD-SecF complex has been directly demonstrated by in vivo cross-linking mass spectrometry.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
|
GO:0005886
plasma membrane
|
IBA
GO_REF:0000033 |
ACCEPT |
Summary: IBA annotation for plasma membrane localization inferred from phylogenetic analysis (PANTHER). SecF is well established as an integral inner membrane protein in bacteria. In P. aeruginosa, the inner membrane corresponds to the plasma membrane. UniProt confirms cell inner membrane localization via HAMAP rule MF_01464.
Reason: SecF is an integral inner membrane protein with six predicted transmembrane helices. Plasma membrane localization is consistent with all structural and functional evidence for SecDF family members across bacteria.
Supporting Evidence:
PMID:21562494
Here, we determined the crystal structure of Thermus thermophilus SecDF at 3.3 Å resolution, revealing a pseudo-symmetrical, 12-helix transmembrane domain belonging to the RND superfamily and two major periplasmic domains, P1 and P4
PMID:7507921
TnphoA fusions have been used to analyze the topology of SecD in the inner membrane; the protein contains six transmembrane stretches and a large periplasmic domain
|
|
GO:0015031
protein transport
|
IBA
GO_REF:0000033 |
KEEP AS NON CORE |
Summary: IBA annotation for involvement in protein transport, inferred from phylogeny. SecF is part of the Sec translocase that mediates protein export across the bacterial inner membrane. This is correct but very general.
Reason: Protein transport is accurate but too broad. The more specific term GO:0043952 (protein transport by the Sec complex) is also annotated and better captures the biological context. This annotation is redundant with the more specific terms.
Supporting Evidence:
PMID:29718185
The bacterial membrane protein SecDF enhances protein translocation across the membrane driven by the complex of SecA ATPase and SecYEG
|
|
GO:0005886
plasma membrane
|
IEA
GO_REF:0000120 |
ACCEPT |
Summary: IEA annotation for plasma membrane from combined automated methods (UniProt subcellular location mapping). Correct and consistent with the IBA annotation above.
Reason: Redundant with the IBA annotation but independently correct. SecF is an integral inner membrane protein confirmed by topology studies and crystal structures.
Supporting Evidence:
PMID:21562494
Here, we determined the crystal structure of Thermus thermophilus SecDF at 3.3 Å resolution, revealing a pseudo-symmetrical, 12-helix transmembrane domain belonging to the RND superfamily and two major periplasmic domains, P1 and P4
|
|
GO:0006605
protein targeting
|
IEA
GO_REF:0000104 |
KEEP AS NON CORE |
Summary: IEA annotation from UniRule for involvement in protein targeting. SecDF enhances the translocation of preproteins across the membrane, which is part of the protein targeting process. However, SecDF functions specifically in the translocation step rather than in the targeting/recognition step per se.
Reason: Protein targeting is a broader process that includes signal recognition and membrane targeting. SecDF specifically functions in the translocation step, after the preprotein has already been targeted to the membrane. This annotation is not incorrect but is less precise than the Sec-complex-specific terms.
Supporting Evidence:
PMID:29718185
SecDF is proposed to undergo repeated conformational transitions to pull out the precursor protein from the SecYEG channel into the periplasm
|
|
GO:0006886
intracellular protein transport
|
IEA
GO_REF:0000002 |
KEEP AS NON CORE |
Summary: IEA annotation from InterPro2GO mapping (IPR005665 SecF bacterial family). SecF is involved in transporting preproteins from the cytoplasm across the inner membrane to the periplasm, which is intracellular protein transport in the context of Gram-negative bacteria.
Reason: Correct but general. The term captures the overall transport function but does not specify the Sec pathway or the PMF-dependent mechanism. More specific terms are available and already annotated.
Supporting Evidence:
PMID:21562494
Protein translocation across the bacterial membrane, mediated by the secretory translocon SecYEG and the SecA ATPase, is enhanced by proton motive force and membrane-integrated SecDF
|
|
GO:0015450
protein-transporting ATPase activity
|
IEA
GO_REF:0000002 |
MODIFY |
Summary: IEA annotation from InterPro2GO mapping (IPR005665). This is an incorrect annotation for SecF. SecF does not have ATPase activity. The ATPase function in the Sec translocase belongs to SecA. SecDF instead uses the proton motive force to drive conformational changes that enhance translocation.
Reason: SecF/SecDF is explicitly not an ATPase. Multiple structural and biochemical studies demonstrate that SecDF uses the proton motive force, not ATP hydrolysis, to enhance translocation. Tsukazaki et al. (2011) identified an ATP-independent translocation step requiring SecDF and PMF. The appropriate MF term is GO:0009977 (proton motive force dependent protein transmembrane transporter activity), as SecDF conducts protons coupled to polypeptide movement. SecF contributes to the translocase complex activity but does not independently enable ATPase activity.
Proposed replacements:
proton motive force dependent protein transmembrane transporter activity
Supporting Evidence:
PMID:21562494
we propose that SecDF functions as a membrane-integrated chaperone, powered by proton motive force, to achieve ATP-independent protein translocation
PMID:29718185
Once SecDF captures the precursor protein on the periplasmic surface, SecDF can complete protein translocation even if SecA function is inactivated by ATP depletion, implying that SecDF is a protein-translocation motor that works independent of SecA
PMID:28467902
SecDF in I form can generate a tunnel that penetrates the transmembrane region and functions as a proton pathway regulated by a conserved Asp residue
|
|
GO:0043952
protein transport by the Sec complex
|
IEA
GO_REF:0000104 |
ACCEPT |
Summary: IEA annotation from UniRule for involvement in protein transport by the Sec complex. This is the most specific and appropriate BP term for SecF function. SecDF is an integral component of the Sec translocase that enhances protein export.
Reason: This is the most specific and accurate BP annotation for SecF. SecDF is a well-characterized component of the Sec protein translocation machinery. Multiple structural and biochemical studies confirm this role across bacterial species.
Supporting Evidence:
PMID:21562494
Protein translocation across the bacterial membrane, mediated by the secretory translocon SecYEG and the SecA ATPase, is enhanced by proton motive force and membrane-integrated SecDF, which associates with SecYEG
PMID:24550475
The SecY/61 complex forms the protein-channel component of the ubiquitous protein secretion and membrane protein insertion apparatus. The bacterial version SecYEG interacts with the highly conserved YidC and SecDF-YajC subcomplex, which facilitates translocation into and across the membrane
|
|
GO:0065002
intracellular protein transmembrane transport
|
IEA
GO_REF:0000104 |
KEEP AS NON CORE |
Summary: IEA annotation from UniRule for intracellular protein transmembrane transport. SecF facilitates the transport of preproteins across the inner membrane, which is intracellular protein transmembrane transport.
Reason: Correct and somewhat more specific than GO:0006886 (intracellular protein transport) in that it specifies transmembrane transport. However, GO:0043952 (protein transport by the Sec complex) is the most informative annotation. This is a valid parent-level annotation that is kept but not core.
Supporting Evidence:
PMID:29718185
Many newly synthesized proteins in the cytoplasm are programmed to be translocated to the periplasm via the narrow channel that is formed in the center of SecYEG
|
|
GO:0031522
cell envelope Sec protein transport complex
|
ISS
PMID:25800553 Probing the protein interaction network of Pseudomonas aerug... |
NEW |
Summary: SecF is a component of the cell envelope Sec protein transport complex. In P. aeruginosa, the SecD-SecF physical interaction has been directly demonstrated by in vivo chemical cross-linking mass spectrometry. The SecDF subcomplex associates with SecYEG to form the Sec translocon, and with YidC to form the holo-translocon. This annotation is supported by deep research findings in secF-deep-research-bioreason-sft.md which correctly identifies SecF as part of the Sec protein secretion complex.
Reason: GO:0031522 (cell envelope Sec protein transport complex) is the most specific CC term for SecF and captures its role as a subunit of the Sec translocase complex. This is supported by direct cross-linking evidence in P. aeruginosa (PMID:25800553) and extensive structural evidence from E. coli and T. thermophilus.
Supporting Evidence:
PMID:25800553
Structures of three membrane proteins, namely, SecD-SecF, OprF, and OprI are predicted using in vivo cross-linked sites
PMID:24550475
The bacterial version SecYEG interacts with the highly conserved YidC and SecDF-YajC subcomplex, which facilitates translocation into and across the membrane
file:PSEAE/secF/secF-deep-research-bioreason-sft.md
[BioReason SFT] A multi-pass inner-membrane component of the bacterial Sec pathway
|
Q: Has SecF or SecDF been directly studied for essentiality or translocation activity in P. aeruginosa, or is all functional evidence transferred from E. coli and T. thermophilus?
Suggested experts: James E Bruce
Q: Does the SecDF complex in P. aeruginosa interact with YidC and participate in membrane protein insertion as part of a holo-translocon, similar to E. coli?
Experiment: Construct a secF deletion mutant in P. aeruginosa PAO1 and measure protein export efficiency (e.g., secretion of alkaline phosphatase or beta-lactamase) in the presence and absence of PMF (using CCCP). Compare translocation rates to wild type and to secD mutants.
Hypothesis: SecF in P. aeruginosa forms a functional SecDF complex that enhances PMF-dependent protein translocation, analogous to E. coli and T. thermophilus.
Experiment: Generate point mutations in the conserved Asp and Arg residues of P. aeruginosa SecF (identified by alignment with T. thermophilus SecDF). Assay protein export and proton conductance in reconstituted proteoliposomes.
Hypothesis: The conserved Asp and Arg residues in the SecF transmembrane region are essential for proton-coupled translocation in P. aeruginosa.
I begin with the InterPro architecture, which is dominated by SecD/SecF family signatures across the entire polypeptide. Two overlapping family-level annotations, IPR005665 (Protein-export membrane protein SecF, bacterial, residues 8–290 and 8–286), and IPR022813 (Protein-export membrane protein SecD/SecF, archaeal and bacterial, residues 9–286), define a multi-pass membrane protein specialized for the Sec pathway. Within this framework, IPR022646 (SecD/SecF conserved site, residues 31–57) and two short family motifs, IPR022645 (bacterial, residues 35–50 and 147–161), plus IPR022645-like motifs at 195–213 and 227–242, mark conserved sequence elements typically embedded in transmembrane helices that mediate specific protein–protein contacts and proton-coupling elements. The C-terminal half is further delineated by IPR055344 (SecD/SecF, C-terminal, bacterial, residues 92–276) and IPR048634 (SecD/SecF, C-terminal, residues 111–287), indicating a structured periplasmic-facing domain that acts as the functional output of the transmembrane core. The ordered layout—N-terminal transmembrane bundle with conserved contact motifs leading into a C-terminal periplasmic domain—creates a proton-motive-force (PMF)-responsive periplasmic hold/fastener module that engages the Sec translocon.
This architecture causes a molecular function centered on protein binding (GO:0005515), because the conserved transmembrane motifs and the C-terminal periplasmic domain are optimized to bind the periplasmic loops of SecY and to capture emerging secretion substrates. The PMF-coupled conformational changes in the transmembrane bundle are transmitted to the C-terminal domain, enabling a “pull” action on polypeptide segments as they exit the SecYEG channel. Thus, the sequence features specify a binding-driven, energy-coupled periplasmic clamp rather than an enzymatic catalyst.
By binding the translocon and secretion substrates and harnessing the PMF to bias forward movement, the protein directly executes protein secretion by the Sec pathway (GO:0009306). The PMF-dependent conformational cycle in the membrane core powers the periplasmic domain to prevent backsliding and to promote completion of translocation, thereby increasing the efficiency of the Sec machinery.
The multi-pass nature and SecD/SecF family identity place the protein in the plasma membrane (GO:0005886) of a bacterium (i.e., the inner membrane), where it assembles with the Sec translocon. Its binding surfaces and family membership indicate stable residence within the cell envelope Sec protein secretion complex (GO:0031522), where it forms a functional unit with SecD and interfaces with SecY and accessory factors.
Mechanistically, the protein likely forms a PMF-gated heterodimer with Preprotein translocase subunit SecD, where SecD stabilizes the transmembrane scaffold and SecF provides the periplasmic fastener that clamps onto the emerging polypeptide. Contacts with Preprotein translocase subunit SecY and its partner YajC organize the translocon’s periplasmic vestibule, while coordination with the membrane protein insertase OxaA/YidC ensures that secretory and membrane proteins are correctly channeled and that insertase activity does not impede translocation. In this model, protonation changes within the transmembrane helices trigger conformational cycles that convert the C-terminal periplasmic domain into a unidirectional clamp, thereby pulling substrates through the Sec channel and enhancing throughput under the SecDF paralog system.
## Functional Summary
A multi-pass inner-membrane component of the bacterial Sec pathway that partners with a small membrane subunit to harness the proton motive force and convert it into mechanical work on emerging secretion substrates. Its transmembrane core couples protonation changes to a periplasmic-facing domain that acts as a fastener, preventing backsliding and actively pulling polypeptides through the Sec translocon. By binding the translocon and substrates, it boosts the efficiency of protein export and coordinates with accessory factors at the membrane to maintain productive secretion.
## UniProt Summary
Part of the SecDF complex, which uses the proton motive force (PMF) to complete protein translocation in the SecYEG preprotein-translocation channel.
## InterPro Domains
- IPR005665: Protein-export membrane protein SecF, bacterial (family) [8-290]
- IPR005665: Protein-export membrane protein SecF, bacterial (family) [8-286]
- IPR022813: Protein-export membrane protein SecD/SecF, archaeal and bacterial (family) [9-286]
- IPR022646: Protein-export membrane protein SecD/SecF/SecDF, conserved site (conserved_site) [31-57]
- IPR022645: Protein-export membrane protein SecD/SecF, bacterial (family) [35-50]
- IPR055344: Protein-export membrane protein SecD/SecF, C-terminal, bacterial (domain) [92-276]
- IPR048634: Protein export membrane protein SecD/SecF, C-terminal (domain) [111-287]
- IPR022645: Protein-export membrane protein SecD/SecF, bacterial (family) [147-161]
- IPR022645: Protein-export membrane protein SecD/SecF, bacterial (family) [195-213]
- IPR022645: Protein-export membrane protein SecD/SecF, bacterial (family) [227-242]
## GO Term Predictions
### Molecular Function
### Biological Process
### Cellular Component
secF encodes the protein translocase subunit SecF (UniProt Q9HXI2) in Pseudomonas aeruginosa PAO1 (ordered locus PA3820). SecF is a multi-pass inner membrane protein that is part of the SecDF accessory complex of the general secretory (Sec) pathway. It enhances protein translocation across the cytoplasmic membrane by harnessing the proton motive force (PMF).
The secF gene was identified as part of the P. aeruginosa PAO1 complete genome sequence PMID:10984043. It is encoded at locus PA3820.
The crystal structure of SecDF from Thermus thermophilus was determined at 3.3 A resolution, revealing a pseudo-symmetrical 12-helix transmembrane domain belonging to the RND superfamily and two major periplasmic domains (P1 and P4) [PMID:21562494 "Structure and function of a membrane component SecDF that enhances protein export", "we determined the crystal structure of Thermus thermophilus SecDF at 3.3 A resolution, revealing a pseudo-symmetrical, 12-helix transmembrane domain belonging to the RND superfamily and two major periplasmic domains, P1 and P4"]. This study showed that SecDF functions as a membrane-integrated chaperone powered by PMF: "we propose that SecDF functions as a membrane-integrated chaperone, powered by proton motive force, to achieve ATP-independent protein translocation" PMID:21562494.
Critically, Tsukazaki et al. (2011) identified an ATP-independent step of translocation requiring both SecDF and PMF, with conserved Asp and Arg residues at the SecD/SecF transmembrane interface playing essential roles in proton and preprotein movement: "Electrophysiological analyses revealed that SecDF conducts protons in a manner dependent on pH and the presence of an unfolded protein, with conserved Asp and Arg residues at the transmembrane interface between SecD and SecF playing essential roles in the movements of protons and preproteins" PMID:21562494.
A review by Tsukazaki (2018) summarized the structural basis for SecDF function: "SecDF is proposed to undergo repeated conformational transitions to pull out the precursor protein from the SecYEG channel into the periplasm. Once SecDF captures the precursor protein on the periplasmic surface, SecDF can complete protein translocation even if SecA function is inactivated by ATP depletion, implying that SecDF is a protein-translocation motor that works independent of SecA" PMID:29718185.
Furukawa et al. (2017) reported I-form structures of SecDF at 2.6-2.8 A resolution, revealing that "SecDF in I form can generate a tunnel that penetrates the transmembrane region and functions as a proton pathway regulated by a conserved Asp residue" PMID:28467902.
The SecDF-YajC subcomplex associates with SecYEG and YidC to form the holo-translocon (HTL). Schulze et al. (2014) showed that the HTL differs from SecYEG alone: "It is more effective in cotranslational insertion of membrane proteins and the posttranslational secretion of a beta-barreled outer-membrane protein driven by SecA and ATP becomes much more dependent on the proton-motive force" PMID:24550475.
Alvira et al. (2020) demonstrated that SecDF's periplasmic domains connect the inner membrane Sec machinery to the outer membrane BAM complex: "the proton-motive force (PMF) across the inner-membrane acts at distinct stages of protein secretion: (1) SecA-driven translocation through SecYEG and (2) communication of conformational changes via SecDF across the periplasm to BAM" PMID:33146611.
Pogliano and Beckwith (1994) characterized the secD operon in E. coli, establishing that it contains yajC, secD, and secF: "in addition to secD and secF, it contains the upstream gene yajC" and that "there are fewer than 30 SecD and SecF molecules per cell" PMID:7507921.
Navare et al. (2015) used in vivo chemical cross-linking mass spectrometry to directly demonstrate SecD-SecF interaction in P. aeruginosa cells: "Structures of three membrane proteins, namely, SecD-SecF, OprF, and OprI are predicted using in vivo cross-linked sites" PMID:25800553. This is one of the few studies providing direct physical evidence for the SecD-SecF complex in P. aeruginosa specifically.
The InterPro2GO mapping (GO_REF:0000002) assigns GO:0015450 (protein-transporting ATPase activity) to SecF. This is problematic because SecF itself is NOT an ATPase. The ATPase activity in the Sec pathway belongs to SecA. SecF/SecDF instead uses the proton motive force for its translocation-enhancing function. A more appropriate MF term would be GO:0009977 (proton motive force dependent protein transmembrane transporter activity), as SecDF conducts protons coupled to polypeptide translocation.
The process annotations (protein transport, protein targeting, protein transport by the Sec complex, intracellular protein transmembrane transport) are all broadly correct. GO:0043952 (protein transport by the Sec complex) is the most specific and informative.
Plasma membrane (GO:0005886) is correct -- SecF is an integral inner membrane protein. However, the more specific complex term GO:0031522 (cell envelope Sec protein transport complex) would also be appropriate.
This is the key distinction for annotation: SecF (as part of SecDF) does not hydrolyze ATP. It uses the proton motive force to drive conformational changes that pull preproteins through the SecYEG channel. The ATPase activity in the Sec translocase belongs to SecA. SecF contributes to the overall translocase complex activity but does not independently enable protein-transporting ATPase activity.
Source: secF-deep-research-bioreason-sft.md
The BioReason SFT functional summary states:
A multi-pass inner-membrane component of the bacterial Sec pathway that partners with a small membrane subunit to harness the proton motive force and convert it into mechanical work on emerging secretion substrates. Its transmembrane core couples protonation changes to a periplasmic-facing domain that acts as a fastener, preventing backsliding and actively pulling polypeptides through the Sec translocon. By binding the translocon and substrates, it boosts the efficiency of protein export and coordinates with accessory factors at the membrane to maintain productive secretion.
This summary is largely accurate and well-aligned with the curated review. The key biological facts are correct:
Inner membrane localization: Correctly identifies SecF as a "multi-pass inner-membrane component," matching the curated GO:0005886 (plasma membrane) annotation and the known six-transmembrane topology.
PMF-driven mechanism: Correctly describes SecF as harnessing "the proton motive force" to "convert it into mechanical work." This matches the curated core function annotation of GO:0009977 (proton motive force dependent protein transmembrane transporter activity) and is well supported by Tsukazaki et al. (2011, PMID:21562494).
Pulling mechanism: The description of "preventing backsliding and actively pulling polypeptides through the Sec translocon" accurately reflects the mechanistic model from structural studies (PMID:29718185, PMID:28467902).
Sec pathway context: Correctly places SecF within the Sec translocation machinery.
Minor issues:
The summary refers to SecF partnering with "a small membrane subunit." SecD is not particularly small -- in E. coli, SecD is 615 aa vs SecF at 323 aa. SecD is actually the larger partner. This is a factual inaccuracy, though minor.
The summary does not mention the specific structural feature of the RND superfamily 12-helix transmembrane domain or the conserved Asp/Arg residues critical for proton conduction, but these are fine-grained structural details that may be beyond the expected scope of a functional summary.
The thinking trace describes SecF function as "protein binding (GO:0005515)" as the molecular function. This is too generic -- per curation guidelines, protein binding is uninformative. The actual molecular function is PMF-dependent protein transmembrane transporter activity (GO:0009977).
What is missing:
The summary does not mention the complex-level organization (SecDF-YajC subcomplex, holo-translocon with YidC) or the recently discovered role of SecDF in inter-membrane communication with BAM for outer membrane protein biogenesis (PMID:33146611). These are important aspects of SecDF biology but may represent advanced knowledge beyond a basic functional summary.
Comparison with interpro2go:
The InterPro2GO annotations for SecF (via GO_REF:0000002, IPR005665) assign:
- GO:0006886 (intracellular protein transport) -- correct but generic
- GO:0015450 (protein-transporting ATPase activity) -- incorrect, as SecF is not an ATPase
BioReason's functional summary significantly outperforms the InterPro2GO MF annotation. Where InterPro2GO incorrectly assigns ATPase activity (conflating SecA's function with SecF's), BioReason correctly identifies the PMF-dependent mechanism. This is a genuine biological insight beyond what InterPro2GO provides, demonstrating that BioReason is not simply recapitulating the automated mapping but is synthesizing information from the InterPro domain architecture to produce a more nuanced functional description.
However, the BioReason thinking trace does propose GO:0005515 (protein binding) as the molecular function, which is similarly uninformative to the InterPro2GO ATPase annotation, albeit for different reasons. The functional summary text itself is more accurate than either the GO term proposed in the trace or the InterPro2GO mapping.
The thinking trace demonstrates solid reasoning from the InterPro domain architecture. It correctly identifies:
- The SecD/SecF family signatures (IPR005665, IPR022813)
- The conserved site motifs (IPR022646, IPR022645)
- The C-terminal periplasmic domain (IPR048634, IPR055344)
- The multi-pass membrane topology
- The PMF-coupling mechanism
The trace's mechanistic model of "protonation changes within the transmembrane helices trigger conformational cycles that convert the C-terminal periplasmic domain into a unidirectional clamp" is a reasonable interpretation consistent with the structural data from Tsukazaki et al. and Furukawa et al.
The trace correctly identifies interaction partners (SecD, SecY, YajC, YidC/OxaA), demonstrating good knowledge of the Sec translocase complex composition.
One weakness: the trace does not distinguish between SecD and SecF contributions to the SecDF heterodimer. In T. thermophilus, SecDF is a single fused polypeptide, but in most bacteria including P. aeruginosa, SecD and SecF are separate proteins. The trace treats the protein as if it has both SecD-like and SecF-like properties, which may partly explain the reference to a "small membrane subunit" partner.
id: Q9HXI2
gene_symbol: secF
product_type: PROTEIN
status: DRAFT
taxon:
id: NCBITaxon:208964
label: Pseudomonas aeruginosa (strain ATCC 15692 / DSM 22644 / CIP 104116 / JCM
14847 / LMG 12228 / 1C / PRS 101 / PAO1)
description: SecF is a multi-pass inner membrane protein that forms the SecDF accessory
complex together with SecD. As part of the Sec protein translocase, SecDF uses the
proton motive force (PMF) to enhance protein translocation across the cytoplasmic
membrane after the ATP-dependent function of SecA. SecDF undergoes PMF-driven conformational
cycles that pull preproteins through the SecYEG channel into the periplasm, functioning
as a membrane-integrated chaperone that works independently of ATP. In P. aeruginosa,
the SecD-SecF complex has been directly demonstrated by in vivo cross-linking mass
spectrometry.
existing_annotations:
- term:
id: GO:0005886
label: plasma membrane
evidence_type: IBA
original_reference_id: GO_REF:0000033
review:
summary: IBA annotation for plasma membrane localization inferred from phylogenetic
analysis (PANTHER). SecF is well established as an integral inner membrane protein
in bacteria. In P. aeruginosa, the inner membrane corresponds to the plasma membrane.
UniProt confirms cell inner membrane localization via HAMAP rule MF_01464.
action: ACCEPT
reason: SecF is an integral inner membrane protein with six predicted transmembrane
helices. Plasma membrane localization is consistent with all structural and functional
evidence for SecDF family members across bacteria.
supported_by:
- reference_id: PMID:21562494
supporting_text: Here, we determined the crystal structure of Thermus thermophilus
SecDF at 3.3 Å resolution, revealing a pseudo-symmetrical, 12-helix transmembrane
domain belonging to the RND superfamily and two major periplasmic domains, P1
and P4
- reference_id: PMID:7507921
supporting_text: TnphoA fusions have been used to analyze the topology of SecD
in the inner membrane; the protein contains six transmembrane stretches and a
large periplasmic domain
- term:
id: GO:0015031
label: protein transport
evidence_type: IBA
original_reference_id: GO_REF:0000033
review:
summary: IBA annotation for involvement in protein transport, inferred from phylogeny.
SecF is part of the Sec translocase that mediates protein export across the
bacterial inner membrane. This is correct but very general.
action: KEEP_AS_NON_CORE
reason: Protein transport is accurate but too broad. The more specific term GO:0043952
(protein transport by the Sec complex) is also annotated and better captures the
biological context. This annotation is redundant with the more specific terms.
supported_by:
- reference_id: PMID:29718185
supporting_text: The bacterial membrane protein SecDF enhances protein translocation
across the membrane driven by the complex of SecA ATPase and SecYEG
- term:
id: GO:0005886
label: plasma membrane
evidence_type: IEA
original_reference_id: GO_REF:0000120
review:
summary: IEA annotation for plasma membrane from combined automated methods (UniProt
subcellular location mapping). Correct and consistent with the IBA annotation above.
action: ACCEPT
reason: Redundant with the IBA annotation but independently correct. SecF is an
integral inner membrane protein confirmed by topology studies and crystal structures.
supported_by:
- reference_id: PMID:21562494
supporting_text: Here, we determined the crystal structure of Thermus thermophilus
SecDF at 3.3 Å resolution, revealing a pseudo-symmetrical, 12-helix transmembrane
domain belonging to the RND superfamily and two major periplasmic domains, P1
and P4
- term:
id: GO:0006605
label: protein targeting
evidence_type: IEA
original_reference_id: GO_REF:0000104
review:
summary: IEA annotation from UniRule for involvement in protein targeting. SecDF
enhances the translocation of preproteins across the membrane, which is part of
the protein targeting process. However, SecDF functions specifically in the
translocation step rather than in the targeting/recognition step per se.
action: KEEP_AS_NON_CORE
reason: Protein targeting is a broader process that includes signal recognition and
membrane targeting. SecDF specifically functions in the translocation step, after
the preprotein has already been targeted to the membrane. This annotation is not
incorrect but is less precise than the Sec-complex-specific terms.
supported_by:
- reference_id: PMID:29718185
supporting_text: SecDF is proposed to undergo repeated conformational transitions
to pull out the precursor protein from the SecYEG channel into the periplasm
- term:
id: GO:0006886
label: intracellular protein transport
evidence_type: IEA
original_reference_id: GO_REF:0000002
review:
summary: IEA annotation from InterPro2GO mapping (IPR005665 SecF bacterial family).
SecF is involved in transporting preproteins from the cytoplasm across the inner
membrane to the periplasm, which is intracellular protein transport in the context
of Gram-negative bacteria.
action: KEEP_AS_NON_CORE
reason: Correct but general. The term captures the overall transport function but
does not specify the Sec pathway or the PMF-dependent mechanism. More specific
terms are available and already annotated.
supported_by:
- reference_id: PMID:21562494
supporting_text: Protein translocation across the bacterial membrane, mediated
by the secretory translocon SecYEG and the SecA ATPase, is enhanced by proton
motive force and membrane-integrated SecDF
- term:
id: GO:0015450
label: protein-transporting ATPase activity
evidence_type: IEA
original_reference_id: GO_REF:0000002
review:
summary: IEA annotation from InterPro2GO mapping (IPR005665). This is an incorrect
annotation for SecF. SecF does not have ATPase activity. The ATPase function in
the Sec translocase belongs to SecA. SecDF instead uses the proton motive force
to drive conformational changes that enhance translocation.
action: MODIFY
reason: SecF/SecDF is explicitly not an ATPase. Multiple structural and biochemical
studies demonstrate that SecDF uses the proton motive force, not ATP hydrolysis,
to enhance translocation. Tsukazaki et al. (2011) identified an ATP-independent
translocation step requiring SecDF and PMF. The appropriate MF term is GO:0009977
(proton motive force dependent protein transmembrane transporter activity), as
SecDF conducts protons coupled to polypeptide movement. SecF contributes to the
translocase complex activity but does not independently enable ATPase activity.
proposed_replacement_terms:
- id: GO:0009977
label: proton motive force dependent protein transmembrane transporter activity
supported_by:
- reference_id: PMID:21562494
supporting_text: we propose that SecDF functions as a membrane-integrated chaperone,
powered by proton motive force, to achieve ATP-independent protein translocation
- reference_id: PMID:29718185
supporting_text: Once SecDF captures the precursor protein on the periplasmic
surface, SecDF can complete protein translocation even if SecA function is
inactivated by ATP depletion, implying that SecDF is a protein-translocation
motor that works independent of SecA
- reference_id: PMID:28467902
supporting_text: SecDF in I form can generate a tunnel that penetrates the
transmembrane region and functions as a proton pathway regulated by a conserved
Asp residue
- term:
id: GO:0043952
label: protein transport by the Sec complex
evidence_type: IEA
original_reference_id: GO_REF:0000104
review:
summary: IEA annotation from UniRule for involvement in protein transport by the
Sec complex. This is the most specific and appropriate BP term for SecF function.
SecDF is an integral component of the Sec translocase that enhances protein export.
action: ACCEPT
reason: This is the most specific and accurate BP annotation for SecF. SecDF is a
well-characterized component of the Sec protein translocation machinery. Multiple
structural and biochemical studies confirm this role across bacterial species.
supported_by:
- reference_id: PMID:21562494
supporting_text: Protein translocation across the bacterial membrane, mediated
by the secretory translocon SecYEG and the SecA ATPase, is enhanced by proton
motive force and membrane-integrated SecDF, which associates with SecYEG
- reference_id: PMID:24550475
supporting_text: The SecY/61 complex forms the protein-channel component of the
ubiquitous protein secretion and membrane protein insertion apparatus. The
bacterial version SecYEG interacts with the highly conserved YidC and
SecDF-YajC subcomplex, which facilitates translocation into and across the
membrane
- term:
id: GO:0065002
label: intracellular protein transmembrane transport
evidence_type: IEA
original_reference_id: GO_REF:0000104
review:
summary: IEA annotation from UniRule for intracellular protein transmembrane
transport. SecF facilitates the transport of preproteins across the inner
membrane, which is intracellular protein transmembrane transport.
action: KEEP_AS_NON_CORE
reason: Correct and somewhat more specific than GO:0006886 (intracellular protein
transport) in that it specifies transmembrane transport. However, GO:0043952
(protein transport by the Sec complex) is the most informative annotation. This
is a valid parent-level annotation that is kept but not core.
supported_by:
- reference_id: PMID:29718185
supporting_text: Many newly synthesized proteins in the cytoplasm are programmed
to be translocated to the periplasm via the narrow channel that is formed in
the center of SecYEG
- term:
id: GO:0031522
label: cell envelope Sec protein transport complex
evidence_type: ISS
original_reference_id: PMID:25800553
review:
summary: SecF is a component of the cell envelope Sec protein transport complex.
In P. aeruginosa, the SecD-SecF physical interaction has been directly demonstrated
by in vivo chemical cross-linking mass spectrometry. The SecDF subcomplex associates
with SecYEG to form the Sec translocon, and with YidC to form the holo-translocon.
This annotation is supported by deep research findings in secF-deep-research-bioreason-sft.md
which correctly identifies SecF as part of the Sec protein secretion complex.
action: NEW
reason: GO:0031522 (cell envelope Sec protein transport complex) is the most
specific CC term for SecF and captures its role as a subunit of the Sec
translocase complex. This is supported by direct cross-linking evidence in P.
aeruginosa (PMID:25800553) and extensive structural evidence from E. coli and T.
thermophilus.
supported_by:
- reference_id: PMID:25800553
supporting_text: Structures of three membrane proteins, namely, SecD-SecF, OprF,
and OprI are predicted using in vivo cross-linked sites
- reference_id: PMID:24550475
supporting_text: The bacterial version SecYEG interacts with the highly conserved
YidC and SecDF-YajC subcomplex, which facilitates translocation into and across
the membrane
- reference_id: file:PSEAE/secF/secF-deep-research-bioreason-sft.md
supporting_text: "[BioReason SFT] A multi-pass inner-membrane component of the\
\ bacterial Sec pathway"
references:
- id: file:PSEAE/secF/secF-deep-research-bioreason-sft.md
title: BioReason SFT deep research for secF
findings:
- statement: BioReason SFT trace correctly identifies SecF as a PMF-driven inner
membrane component of the Sec pathway that uses protonation-driven conformational
changes to pull polypeptides through the translocon.
- id: GO_REF:0000002
title: Gene Ontology annotation through association of InterPro records with GO terms
findings:
- statement: InterPro2GO mapping from IPR005665 (SecF bacterial family) assigns
GO:0006886 (intracellular protein transport) and GO:0015450 (protein-transporting
ATPase activity). The BP annotation is appropriate but the MF annotation is
incorrect -- SecF is not an ATPase.
- id: GO_REF:0000033
title: Annotation inferences using phylogenetic trees
findings:
- statement: PANTHER phylogenetic inference provides IBA annotations for plasma
membrane localization and protein transport, both well supported.
- id: GO_REF:0000104
title: Electronic Gene Ontology annotations created by transferring manual GO
annotations between related proteins based on shared sequence features
findings:
- statement: UniRule transfers provide annotations for protein targeting, protein
transport by the Sec complex, and intracellular protein transmembrane transport,
all appropriate for SecF.
- id: GO_REF:0000120
title: Combined Automated Annotation using Multiple IEA Methods
findings:
- statement: Combined IEA methods confirm plasma membrane localization, consistent
with known SecF inner membrane topology.
- id: PMID:10984043
title: Complete genome sequence of Pseudomonas aeruginosa PAO1, an opportunistic
pathogen
findings:
- statement: Reports the complete genome sequence of P. aeruginosa PAO1, which
includes the secF gene at ordered locus PA3820.
supporting_text: Here we report the complete sequence of P. aeruginosa strain PAO1
- id: PMID:21562494
title: Structure and function of a membrane component SecDF that enhances protein
export
findings:
- statement: Landmark study determining the crystal structure of T. thermophilus
SecDF at 3.3 A, revealing a 12-helix TM domain (RND superfamily) and two
periplasmic domains. Demonstrates SecDF is a PMF-driven membrane-integrated
chaperone for ATP-independent translocation.
supporting_text: we propose that SecDF functions as a membrane-integrated chaperone,
powered by proton motive force, to achieve ATP-independent protein translocation
- statement: Conserved Asp and Arg residues at the SecD/SecF TM interface are
essential for proton conduction and preprotein movement.
supporting_text: Electrophysiological analyses revealed that SecDF conducts protons
in a manner dependent on pH and the presence of an unfolded protein, with
conserved Asp and Arg residues at the transmembrane interface between SecD and
SecF playing essential roles in the movements of protons and preproteins
- id: PMID:29718185
title: Structure-based working model of SecDF, a proton-driven bacterial protein
translocation factor
findings:
- statement: Review of SecDF structural studies. SecDF is a protein-translocation
motor working independently of SecA, powered by the proton gradient.
supporting_text: Once SecDF captures the precursor protein on the periplasmic
surface, SecDF can complete protein translocation even if SecA function is
inactivated by ATP depletion, implying that SecDF is a protein-translocation
motor that works independent of SecA
- id: PMID:28467902
title: Tunnel Formation Inferred from the I-Form Structures of the Proton-Driven
Protein Secretion Motor SecDF
findings:
- statement: I-form crystal structures at 2.6-2.8 A reveal a proton-conducting
tunnel in the SecDF transmembrane region regulated by a conserved Asp residue.
supporting_text: SecDF in I form can generate a tunnel that penetrates the
transmembrane region and functions as a proton pathway regulated by a conserved
Asp residue
- id: PMID:24550475
title: Membrane protein insertion and proton-motive-force-dependent secretion
through the bacterial holo-translocon SecYEG-SecDF-YajC-YidC
findings:
- statement: The holo-translocon (HTL) comprising SecYEG-SecDF-YajC-YidC is more
effective in co-translational membrane protein insertion and makes post-translational
secretion more PMF-dependent.
supporting_text: It is more effective in cotranslational insertion of membrane
proteins and the posttranslational secretion of a beta-barreled outer-membrane
protein driven by SecA and ATP becomes much more dependent on the proton-motive
force
- id: PMID:33146611
title: Inter-membrane association of the Sec and BAM translocons for bacterial
outer-membrane biogenesis
findings:
- statement: SecDF periplasmic domains connect the inner membrane Sec machinery to
the outer membrane BAM complex, enabling inter-membrane communication driven by PMF.
supporting_text: "the proton-motive force (PMF) across the inner-membrane acts at\
\ distinct stages of protein secretion: (1) SecA-driven translocation through SecYEG\
\ and (2) communication of conformational changes via SecDF across the periplasm\
\ to BAM"
- id: PMID:25800553
title: Probing the protein interaction network of Pseudomonas aeruginosa cells by
chemical cross-linking mass spectrometry
findings:
- statement: In vivo cross-linking in P. aeruginosa directly demonstrates the
SecD-SecF physical interaction and provides structural constraints for the
complex.
supporting_text: Structures of three membrane proteins, namely, SecD-SecF, OprF,
and OprI are predicted using in vivo cross-linked sites
- id: PMID:7507921
title: Genetic and molecular characterization of the Escherichia coli secD operon
and its products
findings:
- statement: Characterization of the secD operon in E. coli, establishing that it
contains yajC, secD, and secF. SecD and SecF are present at fewer than 30
molecules per cell.
supporting_text: in addition to secD and secF, it contains the upstream gene yajC
...there are fewer than 30 SecD and SecF molecules per cell
- id: PMID:28636909
title: "Electric-Field-Induced Protein Translocation via a Conformational Transition in SecDF: An MD Study."
findings:
- statement: Molecular dynamics simulations show that the transmembrane electrical
potential lowers the free-energy barrier for the F-form to I-form conformational
transition in SecDF, providing a mechanistic link between PMF and SecDF function.
supporting_text: the interaction of the P1 domain dipole moment with the TM electrical
field considerably lowers the free-energy barrier in the direction of F-form to
I-form transition
core_functions:
- description: SecF, as part of the SecDF complex, functions as a PMF-driven accessory
component of the Sec protein translocase. It uses the proton motive force to power
conformational cycles in its periplasmic domain that pull preproteins through the
SecYEG channel, enhancing protein translocation across the inner membrane independently
of ATP.
molecular_function:
id: GO:0009977
label: proton motive force dependent protein transmembrane transporter activity
contributes_to_molecular_function:
id: GO:0015450
label: protein-transporting ATPase activity
directly_involved_in:
- id: GO:0043952
label: protein transport by the Sec complex
locations:
- id: GO:0005886
label: plasma membrane
in_complex:
id: GO:0031522
label: cell envelope Sec protein transport complex
supported_by:
- reference_id: PMID:21562494
supporting_text: we propose that SecDF functions as a membrane-integrated chaperone,
powered by proton motive force, to achieve ATP-independent protein translocation
- reference_id: PMID:29718185
supporting_text: Once SecDF captures the precursor protein on the periplasmic
surface, SecDF can complete protein translocation even if SecA function is
inactivated by ATP depletion, implying that SecDF is a protein-translocation
motor that works independent of SecA
- reference_id: PMID:25800553
supporting_text: Structures of three membrane proteins, namely, SecD-SecF, OprF,
and OprI are predicted using in vivo cross-linked sites
suggested_questions:
- question: Has SecF or SecDF been directly studied for essentiality or translocation
activity in P. aeruginosa, or is all functional evidence transferred from E. coli
and T. thermophilus?
experts:
- James E Bruce
- question: Does the SecDF complex in P. aeruginosa interact with YidC and participate
in membrane protein insertion as part of a holo-translocon, similar to E. coli?
experts: []
suggested_experiments:
- hypothesis: SecF in P. aeruginosa forms a functional SecDF complex that enhances
PMF-dependent protein translocation, analogous to E. coli and T. thermophilus.
description: Construct a secF deletion mutant in P. aeruginosa PAO1 and measure
protein export efficiency (e.g., secretion of alkaline phosphatase or beta-lactamase)
in the presence and absence of PMF (using CCCP). Compare translocation rates to
wild type and to secD mutants.
- hypothesis: The conserved Asp and Arg residues in the SecF transmembrane region are
essential for proton-coupled translocation in P. aeruginosa.
description: Generate point mutations in the conserved Asp and Arg residues of P.
aeruginosa SecF (identified by alignment with T. thermophilus SecDF). Assay protein
export and proton conductance in reconstituted proteoliposomes.