MlaA (formerly VacJ) is an outer-membrane lipoprotein that forms the outer-membrane interface of the Mla (maintenance of lipid asymmetry) intermembrane phospholipid transport system. After cleavage of its lipoprotein signal peptide and N-terminal lipidation, MlaA folds as a monomeric, mostly alpha-helical, doughnut-shaped protein embedded in the inner (periplasmic) leaflet of the outer membrane, where it associates stably with the trimeric porins OmpC and OmpF. A ridge around its central amphipathic, hydrophilic channel reaches toward the outer leaflet, allowing phospholipids that have become mislocalized in the lipopolysaccharide-rich outer leaflet to enter the channel, while inner-leaflet phospholipids are excluded. A gated beta-hairpin loop and C-terminal helices at the periplasmic opening of the channel recruit and dock the soluble carrier MlaC, juxtaposing the MlaA channel with the MlaC lipid-binding cavity so that phospholipid passes from the outer membrane into MlaC without an energy source at the outer membrane; MlaC then delivers it to the inner-membrane ABC transporter MlaFEDB. Loss of MlaA causes phospholipid accumulation in the outer leaflet, increased outer membrane permeability and sensitivity to SDS-EDTA, whereas the dominant gain-of-function allele mlaA* lets phospholipids flow into the outer leaflet, causing lipopolysaccharide increase, vesiculation and stationary-phase death.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005515 protein binding | IPI PMID:26314242 Osmoporin OmpC forms a complex with MlaA to maintain outer m... | REMOVE | Summary: MlaA interacts specifically with the trimeric porin OmpF (and OmpC), as stated in the abstract of Chong et al. 2015; the MlaA-porin complex is the outer-membrane component of the Mla system. Reason: The interaction itself is well established (also by crystal and cryo-EM structures of MlaA-OmpF/OmpC, PMID:29038444, PMID:38092770), but bare protein binding carries no functional information. The biologically meaningful content (MlaA acting with porins in outer-membrane phospholipid removal) is captured by the cell outer membrane, intermembrane phospholipid transfer and the contributes_to phospholipid transfer activity core function. No more specific porin-binding molecular function term exists that would add information. Removal follows the protein binding policy and does not question the reported interaction. Supporting Evidence: PMID:26314242 We show that the OM lipoprotein MlaA interacts specifically with OmpC and OmpF. |
| GO:0005515 protein binding | IPI PMID:26314242 Osmoporin OmpC forms a complex with MlaA to maintain outer m... | REMOVE | Summary: MlaA interaction with osmoporin OmpC, the porin that functionally partners MlaA in maintaining outer-membrane lipid asymmetry. Reason: As for the OmpF row: the OmpC-MlaA complex is real and functionally important (removing OmpC phenocopies mlaA loss in stationary phase), but generic protein binding is uninformative. The function of the complex is represented by the process and core-function annotations. Removal does not dispute the interaction. Supporting Evidence: PMID:26314242 Removing OmpC, but not OmpF, causes accumulation of PLs in the outer leaflet of the OM in stationary phase, as was previously observed for MlaA. |
| GO:0005515 protein binding | IPI PMID:28388411 Architectures of Lipid Transport Systems for the Bacterial O... | REMOVE | Summary: Ekiert et al. 2017 purified MlaA as a stable complex with OmpF and used it to show that MlaC binds the outer-membrane MlaA-OmpF assembly. Reason: Co-purification of MlaA with OmpF is a genuine physical association, but generic protein binding conveys no function. The informative content of this paper for MlaA (an MlaA-OmpF complex that MlaC docks onto for lipid exchange) is captured by the intermembrane phospholipid transfer process and the contributes_to phospholipid transfer activity core function. Supporting Evidence: PMID:28388411 We over-expressed and purified MlaA from E. coli, which co-purified as a stable complex with OmpF under our culture conditions, as confirmed by mass spectrometry. |
| GO:0009279 cell outer membrane | IEA GO_REF:0000044 | ACCEPT | Summary: MlaA is an outer-membrane lipoprotein embedded in the inner leaflet of the outer membrane in complex with OmpC/OmpF. Reason: Correct and appropriately specific. The outer-membrane location is supported experimentally (porin association that is sufficient to target non-lipidated MlaA to the OM; crystal structure placing MlaA in the inner leaflet of the OM). Supporting Evidence: PMID:26314242 This interaction is sufficient to localize MlaA lacking its lipid anchor to the OM. PMID:29038444 MlaA is a monomeric Ξ±-helical OM protein that functions as a phospholipid translocation channel |
| GO:0009279 cell outer membrane | RCA PMID:12876315 Prediction of lipoprotein signal peptides in Gram-negative b... | ACCEPT | Summary: LipoP-based prediction of a lipoprotein signal peptide; for MlaA the predicted outer-membrane lipoprotein location is confirmed experimentally. Reason: The computational prediction (SPaseII lipoprotein signal peptide, lipidated Cys18) agrees with experimental localization of MlaA to the outer membrane, so the annotation is correct and consistent with the IEA row for the same term. Supporting Evidence: PMID:12876315 The results for Escherichia coli K12 were compared with new experimental data, and the predictions by the HMM agree well with the experimentally verified lipoproteins. PMID:29848551 we established that MlaA interacts extensively with OmpC and is located entirely within the lipid bilayer |
| GO:0016020 membrane | IEA GO_REF:0000002 | MODIFY | Summary: InterPro (MlaA family) mapping to the generic membrane term; correct but far less specific than the known outer-membrane location. Reason: MlaA is specifically an outer-membrane protein, already annotated to GO:0009279 cell outer membrane (ACCEPT above); the informative descendant should replace the generic membrane term. Proposed replacements: cell outer membrane Supporting Evidence: PMID:29038444 forming a ~20-Γ
-thick doughnut embedded in the inner leaflet of the OM with a central, amphipathic pore |
| GO:0120010 intermembrane phospholipid transfer | IBA GO_REF:0000033 | ACCEPT | Summary: PAINT places intermembrane phospholipid transfer at the MlaA family ancestral node; E. coli MlaA is the experimentally characterized member (and appears in its own WITH/FROM as an expected seed). Reason: MlaA directly performs the outer-membrane step of Mla-mediated phospholipid movement between the outer membrane and the inner membrane: it removes phospholipids from the outer leaflet through its channel and hands them to MlaC. This is participation, not mere requirement, so involved_in is appropriate. The term is direction-neutral, which suits the residual debate on anterograde versus retrograde net flux. Supporting Evidence: PMID:38092770 in the first step, the OmpC-MlaA complex transfers PLs to the periplasmic chaperone MlaC, but mechanistic details are lacking. PMID:29038444 The OM lipoprotein MlaA performs the first step in this process via an unknown mechanism that does not require external energy input. |
| GO:0120010 intermembrane phospholipid transfer | IDA PMID:26314242 Osmoporin OmpC forms a complex with MlaA to maintain outer m... | ACCEPT | Summary: Chong et al. 2015 establish that the OmpC-MlaA complex is the outer-membrane component of the Mla system and propose that it removes phospholipids directly from the outer leaflet. Reason: Consistent with the IBA row and with later structural and biochemical work showing MlaA-to-MlaC phospholipid transfer at the outer membrane (PMID:38092770). Only the abstract is cached, but the abstract itself states the role of the OmpC-MlaA complex in the Mla pathway, and the curator-assigned process is clearly correct for MlaA. Supporting Evidence: PMID:26314242 the OmpC-MlaA complex may function to remove PLs directly from the outer leaflet to maintain OM lipid asymmetry PMID:38092770 Using in vivo photo- and disulfide-crosslinking, we demonstrate that MlaC interacts with MlaA directly at its periplasmic base, in a manner that juxtaposes the lipid binding cavity of MlaC with the hydrophilic channel of MlaA. |
Loading supporting contentβ¦
Download this section (compressed HTML)Q: Does MlaA itself bind phospholipid with measurable affinity, or does it act only as a channel through which lipids partition into MlaC? (This decides whether GO:0005543 phospholipid binding should be asserted for MlaA.)
Q: Is there, or should there be, a GO activity term for outer-membrane phospholipid extraction by a membrane-embedded channel that hands lipid to a soluble carrier, distinct from GO:0120014 (which includes carriage through the aqueous phase)?
Q: What role, if any, do OmpC/OmpF play in lipid movement through MlaA beyond positioning and stabilizing it?
Experiment: Reconstitute MlaA (wild type, MlaA*, and the hairpin-loop 3G3P variant) in asymmetric outer-membrane mimetic liposomes with labelled outer-leaflet phospholipid, and measure direct transfer to apo-MlaC with and without OmpC, to quantify MlaA-dependent extraction and its directionality.
Type: in vitro phospholipid transfer assay
Loading supporting contentβ¦
Download this section (compressed HTML)Loading supporting contentβ¦
Download this section (compressed HTML)