PA4178 (eftM, EF-Tu-modifying enzyme) encodes EftM, a 247-residue class I S-adenosyl-L-methionine (SAM)-dependent methyltransferase of the Methyltransf_25 (PF13649) family. Purified EftM directly and exclusively trimethylates lysine 5 of translation elongation factor EF-Tu in a distributive manner. The enzyme is thermolabile and unfolds irreversibly at 37 degrees C, which explains why EF-Tu is trimethylated when P. aeruginosa grows near 25 degrees C but not at 37 degrees C. The G50R substitution in the predicted SAM-binding motif, found in a clinical isolate, abolishes SAM binding and activity. Trimethylated EF-Tu mimics phosphorylcholine, binds the host platelet-activating factor receptor and is associated with increased adhesion to and invasion of respiratory epithelial cells.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0003824 catalytic activity | IEA GO_REF:0000117 | TOO GENERAL | Summary: Root-level catalytic activity from an ARBA rule. True but entailed by the protein-lysine N-methyltransferase activity asserted in this review. Reason: GO:0016279 protein-lysine N-methyltransferase activity (the MODIFY target of the IDA row) is a descendant of catalytic activity, so this row adds nothing. Supporting Evidence: PMID:26677219 An in vitro methyltransferase assay was employed to show that, in the presence of SAM, EftM directly trimethylates EF-Tu. |
| GO:0006744 ubiquinone biosynthetic process | IBA GO_REF:0000033 | REMOVE | Summary: Ubiquinone biosynthesis propagated from the UbiE/COQ5 node PTN001297884. EftM is a characterised protein (EF-Tu lysine 5) methyltransferase, not a quinone C-methyltransferase. Reason: The seeds of PTN001297884 are UbiE/COQ5 orthologs (E. coli UbiE P0A887, human COQ5 Q5HYK3, yeast COQ5 and others). Their function is sound for that clade, but PA4178 has been shown biochemically to methylate a protein substrate, EF-Tu Lys5, exclusively. PA4178 carries no UbiE-type annotation in UniProt (it is classed in PANTHER PTHR43861, trans-aconitate 2-methyltransferase-related, and the TylM1/DesVI family). Placing it under the UbiE node propagates a function it diverged from. Propagation Review Root cause: PROPAGATION BAD Failure modes: FUNCTIONAL DIVERGENCE Sources checked: PANTHER:PTN001297884 · PTN001297884 SUPPORTS SOURCE BUT NOT TARGET UbiE/COQ5 node; its seeds are genuine ubiquinone C-methyltransferases, but PA4178 is an EF-Tu lysine methyltransferase. UniProtKB:P0A887 · E. coli UbiE SUPPORTS SOURCE BUT NOT TARGET Sound for UbiE; PAO1 has its own UbiE ortholog. Supporting Evidence: PMID:26677219 Mass spectrometry analysis of the in vitro methyltransferase reaction products revealed that EftM exclusively methylates at lysine 5 of EF-Tu in a distributive manner. file:PSEAE/PA4178/PA4178-uniprot.txt PANTHER; PTHR43861; TRANS-ACONITATE 2-METHYLTRANSFERASE-RELATED; 1. |
| GO:0008168 methyltransferase activity | IDA PMID:26677219 Pseudomonas aeruginosa EftM Is a Thermoregulated Methyltrans... | MODIFY | Summary: PseudoCAP IDA and one of ten seeds of the PTN000010968 methyltransferase IBD. Purified EftM trimethylates EF-Tu with SAM as donor, so the term is correct, but the paper supports the more specific protein-lysine N-methyltransferase activity. Reason: The full text shows SAM-dependent trimethylation of EF-Tu lysine 5 by purified enzyme, with the residue identified by mass spectrometry. That is GO:0016279 protein-lysine N-methyltransferase activity, an is_a descendant of methyltransferase activity. As an IBD seed the row is sound: the descendant is valid evidence for the generic node term. Proposed replacements: protein-lysine N-methyltransferase activity Supporting Evidence: PMID:26677219 An in vitro methyltransferase assay was employed to show that, in the presence of SAM, EftM directly trimethylates EF-Tu. PMID:26677219 Mass spectrometry analysis of the in vitro methyltransferase reaction products revealed that EftM exclusively methylates at lysine 5 of EF-Tu in a distributive manner. |
| GO:0008425 2-methoxy-6-polyprenyl-1,4-benzoquinol methyltransferase activity | IBA GO_REF:0000033 | REMOVE | Summary: UbiE/COQ5 C-methyltransferase activity propagated from PTN001297884. EftM methylates a protein lysine instead. Reason: Same over-propagation as the ubiquinone biosynthesis IBA row (see the GO_CENTRAL defect recorded there). The demonstrated activity is an N-methylation of EF-Tu Lys5, not C-methylation of a benzoquinol. Propagation Review Root cause: PROPAGATION BAD Failure modes: FUNCTIONAL DIVERGENCE Sources checked: PANTHER:PTN001297884 · PTN001297884 SUPPORTS SOURCE BUT NOT TARGET Sound for UbiE/COQ5; not for EftM. The defect is recorded once, on the GO:0006744 row. Supporting Evidence: PMID:26677219 Mass spectrometry analysis of the in vitro methyltransferase reaction products revealed that EftM exclusively methylates at lysine 5 of EF-Tu in a distributive manner. |
| GO:0017000 antibiotic biosynthetic process | IEA GO_REF:0000117 | REMOVE | Summary: ARBA association rule, apparently driven by the TylM1/DesVI (macrolide sugar N-methyltransferase) family assignment. EftM methylates EF-Tu and has no known role in antibiotic synthesis. Reason: The only characterised substrate is a protein (EF-Tu Lys5), and no antibiotic or secondary-metabolite product is associated with PA4178. The rule over-propagates a family-level pathway. Supporting Evidence: PMID:26677219 EftM is a thermolabile, SAM-dependent methyltransferase that directly trimethylates lysine 5 of EF-Tu in P. aeruginosa. |
| GO:0018023 peptidyl-lysine trimethylation | IDA PMID:26677219 Pseudomonas aeruginosa EftM Is a Thermoregulated Methyltrans... | NEW | Summary: Proposed process. EftM itself catalyses trimethylation of EF-Tu Lys5 in vitro, and its deletion abolishes the modification in vivo. Reason: EftM performs the step (it is the enzyme, not a substrate or regulator). Mass spectrometry shows trimethyl-lysine 5 formed by purified enzyme. Supporting Evidence: PMID:26677219 Collectively, our results show that EftM is a thermolabile, SAM-dependent methyltransferase that directly trimethylates lysine 5 of EF-Tu in P. aeruginosa. |
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Download this section (compressed HTML)Q: Does PANTHER place PA4178 (EftM) inside the UbiE/COQ5 clade PTN001297884 by mistake, or has EftM diverged from a UbiE-like ancestor so that a loss (NOT) annotation is needed on its branch?
Q: Is a dedicated GO term for EF-Tu lysine N-methyltransferase activity (analogous to the cytochrome c and calmodulin lysine methyltransferase terms) warranted?
Experiment: Determine what fraction of native EF-Tu is trimethylated at 25 and 37 degrees C and whether the modification affects translation elongation or EF-Tu export to the cell surface.
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