IadE (Vapar_1494) is the 162-amino-acid structural beta subunit of the heterohexameric (alpha3beta3) indole-3-acetic acid (IAA) Rieske non-heme iron aromatic-ring-hydroxylating dioxygenase complex (IadD/IadE) in Variovorax paradoxus S110. IadE adopts a single-domain NTF2-like twisted seven-stranded beta-sheet fold with three alpha-helices and trimerizes to form the structural stem base of the mushroom-shaped IadD/IadE heterohexamer, scaffolding the catalytic IadD alpha-subunit trimer cap. Co-expression of iadE with iadD is required for assembly of the functional IadD/IadE dioxygenase holoenzyme and for aerobic IAA catabolism and reversion of auxin-induced root growth inhibition in the plant rhizosphere.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0019380 3-phenylpropionate catabolic process | IEA GO_REF:0000118 | REMOVE | Summary: TreeGrafter over-propagated 3-phenylpropionate catabolic process from distant aromatic dioxygenase beta-subunit paralogs in PTHR41534; IadE specifically functions in indole-3-acetic acid (IAA) catabolism. Reason: Direct genetic, metabolomic, and structural characterization of Variovorax paradoxus S110 IadD/IadE (PMID:36266335, PMID:37459330) establishes that IadE is the structural beta subunit of the indole-3-acetic acid (IAA) dioxygenase complex (IadD/IadE), not a 3-phenylpropionate dioxygenase. Supporting Evidence: PMID:36266335 To test the necessity of these genes, we constructed knockout strains CL14 ΔiadDE and CL14 ΔiadCDE. Both lacked the ability to degrade IAA (Fig. 1d), demonstrating that iadD and iadE are necessary and sufficient for IAA degradation by Variovorax. PMID:37459330 Together, these data suggest that IAA was converted to oxIAA by the dioxygenase system composed of the IadD/E dioxygenase and the IadC reductase (Fig 5E). |
| GO:0016708 oxidoreductase activity, acting on paired donors, with incorporation or reduction of molecular oxygen, NAD(P)H as one donor, and incorporation of two atoms of oxygen into one donor | IDA PMID:37459330 Structural and biochemical characterization of the key compo... | NEW | Summary: IadE contributes to the aromatic-ring-hydroxylating dioxygenase activity of the alpha3beta3 IadD/IadE holoenzyme as its obligate structural beta subunit. Reason: While the active site resides in IadD, IadE is required to assemble the functional alpha3beta3 IadD/IadE dioxygenase complex that converts IAA to oxIAA in the presence of IadC and NADH, and therefore contributes_to GO:0016708. Supporting Evidence: PMID:37459330 IadE is a single-domain protein, containing a twisted seven-stranded β-sheet and 3 α-helices, which may mainly serve as a scaffold protein contributing to proper assembly of the oligomeric enzyme complex. PMID:36266335 To test the necessity of these genes, we constructed knockout strains CL14 ΔiadDE and CL14 ΔiadCDE. Both lacked the ability to degrade IAA (Fig. 1d), demonstrating that iadD and iadE are necessary and sufficient for IAA degradation by Variovorax. |
| GO:0042437 indoleacetic acid catabolic process | IMP PMID:36266335 Diverse MarR bacterial regulators of auxin catabolism in the... | NEW | Summary: IadE is required for indole-3-acetic acid (IAA) catabolism in Variovorax. Reason: Deletion of iadDE abolishes IAA degradation and heterologous expression of iadDE in E. coli confers IAA degradation activity. Supporting Evidence: PMID:36266335 To test the necessity of these genes, we constructed knockout strains CL14 ΔiadDE and CL14 ΔiadCDE. Both lacked the ability to degrade IAA (Fig. 1d), demonstrating that iadD and iadE are necessary and sufficient for IAA degradation by Variovorax. |
| GO:0005829 cytosol | IC PMID:37459330 Structural and biochemical characterization of the key compo... | NEW | Summary: IadE functions in the bacterial cytosol as the beta-subunit trimer base of the soluble IadD/IadE complex. Reason: Inferred by curator (IC) because IadE is a soluble cytoplasmic subunit lacking signal peptides or transmembrane helices that forms the stem of the cytosolic alpha3beta3 IadD/IadE dioxygenase complex. Supporting Evidence: PMID:37459330 An overall mushroom-shaped heterohexamer composed of 3 IadE at the “stem” and 3 IadD in the “cap” was resolved (Fig 3C). |
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Download this section (compressed HTML)Q: Does the IadE trimer stem allosterically modulate substrate binding or product release in the overlying IadD catalytic cap, or is its role purely assembly-scaffolding?
Experiment: Assess the thermal stability, oligomeric state, and iron-center occupancy of recombinant V. paradoxus S110 IadD expressed with and without IadE.
Hypothesis: IadE trimerization templates the assembly of the IadD trimer cap and stabilizes the inter-subunit Rieske-to-mononuclear-iron interface.
Type: Size-exclusion chromatography-MALS and differential scanning fluorimetry
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