PA5089 is PldB (also called Tle5b), one of the two phospholipase D-family type VI secretion system effectors of P. aeruginosa PAO1. It is a 745-residue protein of the PLD/transphosphatidylase superfamily carrying an HxKxxxxD (HKD) catalytic motif, and it is encoded immediately downstream of vgrG5 (PA5090) and upstream of a cluster of three cognate immunity proteins (tli5b1-3, PA5088-PA5086). PldB is exported by a type VI secretion system in a VgrG5-dependent manner and delivered into the periplasm of competing bacteria, where its phospholipase activity is toxic; the producing cell is protected by the Tli5b immunity proteins. PldB has also been reported to promote invasion of eukaryotic host cells, making it a trans-kingdom effector. Its action is therefore an attack on the membrane phospholipids of another cell rather than a step in the bacterium's own lipid metabolism.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0004630 D-type glycerophospholipase activity | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from the PAINT IBD on PTN004499769, the PLD superfamily node. The IBD rests on 24 seeds, among them human PLD1 (Q13393), yeast SPO14 (S000001739), rat Pld1/Pld2 (RGD:3349, RGD:3350), ten Arabidopsis PLDs and the two PAO1 effectors. PA5089 carries the family's HKD motif with three PROSITE-predicted active-site residues, so it sits properly inside the clade. The appearance of Q9HU94 in WITH/FROM reflects its own PseudoCAP experimental row and is expected. Reason: The phylogenetic claim is sound and well seeded by characterised eukaryotic PLDs; D-type glycerophospholipase activity is the defining activity of the family and of the Tle5 effectors. Supporting Evidence: PMID:23552891 Four of the families (Tle1-4) exhibit the GxSxG motif common in esterases and many lipases, while the fifth (Tle5) possesses dual HxKxxxxD motifs found in PLD enzymes |
| GO:0009395 phospholipid catabolic process | IBA GO_REF:0000033 | MARK AS OVER ANNOTATED | Summary: A second IBD on the same node, seeded only by plant and rat PLDs (AT1G55180, AT3G05630, Pld1, Pld2) that hydrolyse phospholipids of their own membranes in lipid turnover and signalling. PldB is a secreted T6SS toxin, hydrolysing phospholipids in the periplasm of a target cell, and nothing indicates that it contributes to phospholipid catabolism in P. aeruginosa itself. Reason: The chemistry is phospholipid hydrolysis, so the term is not simply false, but the process claim transfers the endogenous lipid-turnover role of the eukaryotic seeds onto an interbacterial toxin. The informative process for this gene is delivery of a toxin into a competitor, not degradation of the organism's own phospholipids. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: ROLE CONFLATION CONTEXT OR TISSUE MISMATCH Sources checked: PANTHER:PTN004499769 · PTN004499769 SUPPORTS SOURCE BUT NOT TARGET The GO:0009395 IBD has four seeds, all eukaryotic PLDs acting on their own membrane lipids. The node is wide enough to include secreted bacterial toxins, for which the process reading does not hold. Supporting Evidence: PMID:24832454 PldB targets the periplasm of prokaryotic cells and exerts an antibacterial activity. |
| GO:0003824 catalytic activity | IEA GO_REF:0000002 | TOO GENERAL | Summary: InterPro2GO root-level catalytic activity from the PLD/transphosphatidylase signature. True, but entailed by the accepted GO:0004630. Reason: Root-of-branch term; the specific D-type glycerophospholipase activity asserted elsewhere in this review already implies it. |
| GO:0004630 D-type glycerophospholipase activity | IEA GO_REF:0000003 | ACCEPT | Summary: EC 3.1.4.4 to GO mapping. Consistent with the family assignment and with the experimental row below. Reason: The EC mapping agrees with the HKD-motif family assignment and with the PseudoCAP experimental annotation on the same term. |
| GO:0004630 D-type glycerophospholipase activity | IMP PMID:24832454 A Pseudomonas aeruginosa type VI secretion phospholipase D e... | ACCEPT | Summary: PseudoCAP row, and one of the 24 seeds of the PTN004499769 IBD. The cited paper is about this gene - PA5089 is pldB, which the strain table of PMID:31417515 confirms independently - and describes it as a phospholipase D effector whose delivery into the prey periplasm is antibacterial. Only the abstract is cached, so whether a phospholipase assay underlies the assignment could not be checked; the family's HKD motif and the dedicated immunity proteins both point the same way. Reason: Right gene, right family, and the row cites the paper that characterised this effector. The seed is sound for the node term, which is the family-level activity. Supporting Evidence: PMID:24832454 Here we describe the characterization of a P. aeruginosa H3-T6SS-dependent phospholipase D effector, PldB, and its three tightly linked cognate immunity proteins. |
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Download this section (compressed HTML)Q: Has purified PldB been shown to hydrolyse phospholipids in vitro, and with what head-group specificity, as was done for PldA (phosphatidylcholine and phosphatidylethanolamine)?
Q: PldB carries a single PROSITE-detected HKD motif whereas PldA carries two. Does PldB act as a homodimer that assembles a composite active site?
Q: Should GO offer a process term for delivery of an antibacterial effector into a target cell, so that T6SS toxins are not annotated to the endogenous metabolic process matching their chemistry (here phospholipid catabolism)?
Experiment: Assay purified PldB against defined phospholipid substrates (phosphatidylethanolamine, phosphatidylcholine, cardiolipin), with and without mutation of the predicted active-site residues at positions 642, 644 and 649, and quantify the products by mass spectrometry as was done for PldA.
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