FAD-dependent oxidoreductase Fox (VPARA_05580, A0A0H2M811; EC 1.-.-.-) in Variovorax paradoxus strain TBEA6 is a 474-amino-acid cytosolic flavoprotein of the PCMH/vanillyl-alcohol oxidase type-4 FAD-binding oxidoreductase family (PROSITE PS51387, Pfam PF01565, Pfam PF02913, InterPro IPR016166, InterPro IPR004113) encoded on genomic contig JZWI01000003 (EMBL KLN58443.1) two open reading frames upstream of mdo (VPARA_05600, KLN58445.1, 3-mercaptopropionate dioxygenase) and ~13 open reading frames from act (VPARA_05450, KLN58430.1, 3-sulfinopropionate CoA-transferase) and acd (VPARA_05440, KLN58429.1, 3-sulfinopropionyl-CoA desulfinase). Fox comprises an N-terminal PCMH-type FAD-binding domain (PROSITE PS51387, Pfam PF01565, residues 36-217) and a C-terminal FAD-linked oxidase substrate-binding domain (Pfam PF02913, InterPro IPR004113); its genomic proximity to mdo suggests a candidate role in oxidative organosulfur or hydroxy-acid metabolism, although its physiological substrate has not been biochemically established.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0003824 catalytic activity | IEA GO_REF:0000002 | MODIFY | Summary: Correct but general root molecular function; refine to GO:0016491 (oxidoreductase activity). Reason: Fox (VPARA_05580, EC 1.-.-.-) contains a canonical N-terminal PCMH-type FAD-binding domain (PROSITE PS51387, Pfam PF01565, residues 36-217) and a C-terminal FAD-linked oxidase domain (Pfam PF02913, InterPro IPR004113, InterPro IPR016164); refine root GO:0003824 to GO:0016491 (oxidoreductase activity) rather than the narrower child GO:0016614 (oxidoreductase activity, acting on CH-OH group of donors), because the physiological donor group oxidized by VPARA_05580 in V. paradoxus TBEA6 has not been experimentally determined. Proposed replacements: oxidoreductase activity Supporting Evidence: file:VARPD/fox/fox-uniprot.txt DR Pfam; PF02913; FAD-oxidase_C; 1. DR Pfam; PF01565; FAD_binding_4; 1. |
| GO:0022904 respiratory electron transport chain | IEA GO_REF:0000118 | REMOVE | Summary: TreeGrafter over-propagation from eukaryotic mitochondrial D-lactate / D-2-hydroxyglutarate dehydrogenases (PANTHER PTHR43716); Fox is a catabolic flavoprotein oxidoreductase in the V. paradoxus TBEA6 organosulfur (TDP/3MP) degradation cluster. Reason: TreeGrafter transferred GO:0022904 from PANTHER node PTN008117839 within PTHR43716 (mitochondrial D-2-hydroxyglutarate / D-lactate dehydrogenase). In Variovorax paradoxus TBEA6, fox (VPARA_05580, KLN58443.1) is a soluble flavoprotein oxidoreductase encoded in the 3,3'-thiodipropionic acid (TDP) catabolic gene cluster two ORFs upstream of 3-mercaptopropionate dioxygenase mdo (VPARA_05600, KLN58445.1; PMID:19001372), and is not a respiratory electron transport chain subunit. Supporting Evidence: file:VARPD/fox/fox-uniprot.txt DR PANTHER; PTHR43716:SF2; BLL6224 PROTEIN; 1. DR PANTHER; PTHR43716; D-2-HYDROXYGLUTARATE DEHYDROGENASE, MITOCHONDRIAL; 1. DR Pfam; PF02913; FAD-oxidase_C; 1. DR Pfam; PF01565; FAD_binding_4; 1. |
| GO:0050660 flavin adenine dinucleotide binding | IEA GO_REF:0000002 | TOO GENERAL | Summary: True but uninformative parent molecular function already entailed by the accepted specific child term GO:0071949 (FAD binding). Reason: GO:0050660 (flavin adenine dinucleotide binding) is the direct parent of GO:0071949 (FAD binding) already accepted in this review. Supporting Evidence: file:VARPD/fox/fox-uniprot.txt FT DOMAIN 36..217 FT /note="FAD-binding PCMH-type" FT /evidence="ECO:0000259|PROSITE:PS51387" |
| GO:0071949 FAD binding | IEA GO_REF:0000002 | ACCEPT | Summary: Core cofactor-binding activity of the N-terminal PCMH-type FAD-binding domain (PROSITE PS51387, Pfam PF01565, InterPro IPR016166, residues 36-217). Reason: Fox contains a well-defined N-terminal PCMH-type FAD-binding domain (PROSITE PS51387, Pfam PF01565, InterPro IPR016166, residues 36-217) that coordinates the FAD cofactor required for flavoprotein oxidoreductase catalysis. Supporting Evidence: file:VARPD/fox/fox-uniprot.txt FT DOMAIN 36..217 FT /note="FAD-binding PCMH-type" FT /evidence="ECO:0000259|PROSITE:PS51387" |
| GO:0005829 cytosol | IC file:VARPD/fox/fox-uniprot.txt | NEW | Summary: Core subcellular localization of the soluble 474-aa cytoplasmic flavoprotein Fox. Reason: Inferred by curator (IC) from the 474-aa sequence in file:VARPD/fox/fox-uniprot.txt, which lacks transmembrane helices or secretory signal peptides. Supporting Evidence: file:VARPD/fox/fox-uniprot.txt |
| GO:0016491 oxidoreductase activity | IC file:VARPD/fox/fox-uniprot.txt | NEW | Summary: Core catalytic activity of the FAD-dependent oxidoreductase Fox (EC 1.-.-.-). Reason: Inferred by curator (IC) from the PCMH-type FAD-binding domain (PROSITE PS51387, Pfam PF01565) and C-terminal FAD-linked oxidase domain (Pfam PF02913, InterPro IPR004113) in file:VARPD/fox/fox-uniprot.txt; kept at GO:0016491 rather than GO:0016614 because the physiological substrate of VPARA_05580 has not been experimentally identified. Supporting Evidence: file:VARPD/fox/fox-uniprot.txt DR Pfam; PF02913; FAD-oxidase_C; 1. DR Pfam; PF01565; FAD_binding_4; 1. |
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Download this section (compressed HTML)Q: Does purified Variovorax paradoxus Fox (VPARA_05580) directly catalyze the FAD-dependent oxidative cleavage of 3,3'-thiodipropionic acid (TDP) to 3-mercaptopropionate and 3-oxopropionate/3-hydroxypropionate in vitro?
Suggested experts: Steinbuchel A, Wubbeler JH
Experiment: Heterologously express and purify recombinant VPARA_05580 (Fox) from E. coli, verify noncovalent or covalent FAD cofactor occupancy by UV-Vis spectroscopy, and assay oxidative cleavage of 3,3'-thiodipropionic acid (TDP) to 3-mercaptopropionate by HPLC-ESI-MS.
Hypothesis: Fox catalyzes the initial FAD-dependent oxidative cleavage of TDP into 3-mercaptopropionate for subsequent dioxygenation by Mdo (VPARA_05600).
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