PqqD (PqqA binding protein, VPARA_00930, A0A0H2M992) in Variovorax paradoxus is a 98-amino-acid cofactor-less RiPP recognition element (RRE) peptide chaperone (HAMAP MF_00655, InterPro IPR008792, InterPro IPR022479, NCBIfam TIGR03859, Pfam PF05402) encoded between pqqC (VPARA_00940) and pqqE (VPARA_00920) in the pqqBCDE gene cluster. PqqD binds the unmodified precursor peptide PqqA with nanomolar affinity and forms a ternary PqqA-PqqD-PqqE complex that presents the conserved glutamate and tyrosine residues of PqqA to the active site of the radical SAM cyclase PqqE for initiating carbon-carbon cross-link formation in pyrroloquinoline quinone (PQQ) biosynthesis.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0018189 pyrroloquinoline quinone biosynthetic process | IEA GO_REF:0000120 | ACCEPT | Reason: PqqD (VPARA_00930, A0A0H2M992) is an essential peptide chaperone in the pyrroloquinoline quinone biosynthetic pathway that binds the precursor peptide PqqA and presents it to PqqE for Glu-Tyr cross-linking (PMID:25817994, PMID:26961875). Supporting Evidence: UniProtKB:A0A0H2M992 CC -!- FUNCTION: Functions as a PqqA binding protein and presents PqqA to CC PqqE, in the pyrroloquinoline quinone (PQQ) biosynthetic pathway. CC {ECO:0000256|HAMAP-Rule:MF_00655}. PMID:26961875 These results indicate that PqqE, in conjunction with PqqD, carries out the first step in PQQ biosynthesis |
| GO:0048038 quinone binding | IEA GO_REF:0000002 | REMOVE | Reason: Erroneous InterPro2GO mapping on IPR022479 (PqqD_bac) that conflates participation in the pyrroloquinoline quinone biosynthetic pathway with binding of mature quinones. Biophysical and biochemical studies (PMID:25817994, PMID:32731194) proved that PqqD is a cofactor-less RiPP recognition element (RRE) protein that binds the unmodified precursor peptide PqqA (Kd ~ 200 nM), not a quinone (matching the removal of GO:0048038 in PSEPK/pqqD1). Supporting Evidence: PMID:32731194 The missing factor has turned out to be PqqD, a small, cofactor-less protein that primarily exists as a free protein (and occasionally as a C-terminal fusion protein of PqqC or an N-terminal fusion of PqqE) (Figure 1D).[12] PMID:25817994 Moreover, using SPR and isothermal calorimetry techniques, we establish for the first time that MePqqD binds MePqqA tightly (KD βΌ200 nm). |
| GO:0042277 peptide binding | ISS PMID:25817994 PqqD is a novel peptide chaperone that forms a ternary compl... | NEW | Reason: PqqD functions specifically as a PqqA peptide-binding protein (RiPP recognition element) that binds the leader region of the PqqA precursor peptide with nanomolar affinity (Kd ~ 200 nM) to present it to PqqE (PMID:25817994, UniProtKB:A0A0H2M992). Supporting Evidence: PMID:25817994 Moreover, using SPR and isothermal calorimetry techniques, we establish for the first time that MePqqD binds MePqqA tightly (KD βΌ200 nm). UniProtKB:A0A0H2M992 CC -!- FUNCTION: Functions as a PqqA binding protein and presents PqqA to CC PqqE, in the pyrroloquinoline quinone (PQQ) biosynthetic pathway. CC {ECO:0000256|HAMAP-Rule:MF_00655}. |
| GO:0005829 cytosol | IC UniProtKB:A0A0H2M992 | NEW | Reason: Inferred by curator (IC) because V. paradoxus PqqD (A0A0H2M992) is a soluble 98-residue monomeric protein lacking signal peptides or transmembrane domains that forms a cytosolic PqqA-PqqD-PqqE ternary complex. Supporting Evidence: UniProtKB:A0A0H2M992 CC -!- SUBUNIT: Monomer. Interacts with PqqE. {ECO:0000256|ARBA:ARBA00011741, CC ECO:0000256|HAMAP-Rule:MF_00655}. |
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Download this section (compressed HTML)Q: Which residues of the winged-helix-turn-helix fold in Variovorax paradoxus PqqD (VPARA_00930) form the beta-sheet augmentation interface with the N-terminal leader sequence of PqqA versus the docking interface with PqqE (VPARA_00920)?
Experiment: Measure the binding affinity (Kd) and stoichiometry of purified Variovorax paradoxus PqqD (A0A0H2M992) for synthetic V. paradoxus PqqA peptide and PqqE (A0A0H2MPV4) by isothermal titration calorimetry (ITC) and native mass spectrometry.
Hypothesis: V. paradoxus PqqD binds the N-terminal leader motif of PqqA with sub-micromolar affinity and assembles into a 1:1:1 PqqA-PqqD-PqqE ternary complex required for Glu-Tyr cross-linking.
Type: Isothermal titration calorimetry and native mass spectrometry
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