CYP2U1 is a membrane-associated, heme-containing cytochrome P450 expressed in the thymus and brain and localized to the endoplasmic reticulum and mitochondria. It uses NADPH-derived electrons delivered by cytochrome P450 reductase to hydroxylate long-chain fatty acids, forming 19- and 20-HETE from arachidonic acid. It also oxidizes the indole ring of N-arachidonoylserotonin, and recombinant enzyme can metabolize sorafenib and sunitinib. Biallelic loss of function causes hereditary spastic paraplegia 56; patient and mouse studies associate deficiency with disturbed mitochondrial physiology, lipid balance and folate-related metabolism.
Definition: Catalysis of the reaction: N-arachidonoylserotonin + reduced NADPH–hemoprotein reductase + O2 = 2-oxo-N-arachidonoylserotonin + oxidized NADPH–hemoprotein reductase + H2O + H+. Oxygen is incorporated at indole C2.
Justification: Human CYP2U1 forms the structurally characterized 2-oxo product, represented by literal RHEA:50296 on the official protein record. Bounded current GO/OLS substrate and reaction searches did not identify a precise activity term. GO:0036190 instead describes free indole to indolin-2-one and cannot be substituted for the lipidated-serotonin reaction. This requests a specific term without asserting exhaustive ontology absence or a new physiological process.
Mappings:
Supporting Evidence:
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0004497 monooxygenase activity | IEA GO_REF:0000120 | TOO GENERAL | Summary: Monooxygenase activity is correct but less informative than the resolved catalytic activities. Reason: The current InterPro mappings and matching ARBA00028788 branch 6 support P450 monooxygenase chemistry. Direct human enzyme experiments resolve long-chain fatty-acid omega and omega-1 hydroxylation and a separate N-arachidonoylserotonin oxidation; the general parent adds no distinct catalytic unit. |
| GO:0004497 monooxygenase activity | TAS Reactome:R-HSA-211960 | MODIFY | Summary: The modeled arachidonate reaction is long-chain fatty-acid omega-1 hydroxylation. Reason: The exact Reactome 211960 catalyst path reaches Q7Z449 and forms 19-HETE from C20 arachidonate. This is omega-1 hydroxylation, despite the summation calling its 19-position example omega hydroxylation. Proposed replacements: long-chain fatty acid omega-1 hydroxylase activity |
| GO:0004497 monooxygenase activity | TAS Reactome:R-HSA-2161814 | MODIFY | Summary: Refine the 19-HETE reaction to long-chain fatty-acid omega-1 hydroxylase activity. Reason: The actual CYP(2) catalyst set contains CYP2U1 through entity 2142802/reference 409441. Its event explicitly forms 19-HETE, and recombinant human CYP2U1 independently performs that chemistry. Membership does not assign every substrate specificity of the other set members. Proposed replacements: long-chain fatty acid omega-1 hydroxylase activity |
| GO:0004497 monooxygenase activity | TAS Reactome:R-HSA-2161940 | MODIFY | Summary: Refine the 20-HETE reaction to long-chain fatty-acid omega-hydroxylase activity. Reason: The actual CYP(3) catalyst set contains Q7Z449 and the event forms 20-HETE. Human recombinant turnover supports the specific terminal hydroxylation term already represented by direct annotations. Proposed replacements: long-chain fatty acid omega-hydroxylase activity |
| GO:0004497 monooxygenase activity | TAS Reactome:R-HSA-5602242 | TOO GENERAL | Summary: The gene has monooxygenase activity, but this disease event is not a positive mutant-activity assay. Reason: Reactome 5602242 follows a set of five defective CYP2U1 variants to Q7Z449 and describes failed arachidonate hydroxylation. The positive gene-level parent is independently established by wild-type enzyme studies and is covered more precisely elsewhere. The modeled loss-of-function variants are not evidence of active enzymes. |
| GO:0005506 iron ion binding | IEA GO_REF:0000002 | ACCEPT | Summary: Heme-associated iron binding is integral to CYP2U1 catalysis. Reason: The five literal InterPro records map to the current official CYP2U1 features and include the heme-binding conserved site. Heme-dependent recombinant P450 spectra and oxygenation assays support the cofactor role; this does not demonstrate activity of the severely shortened splice product. |
| GO:0005737 cytoplasm | IBA GO_REF:0000033 | TOO GENERAL | Summary: The cytoplasmic placement is valid but is more broadly scoped than the established membrane localization. Reason: The exact PTN000670347 IBD is on the verified human path. Cytoplasm includes cytoplasmic organelles; this is not a claim of soluble cytosolic CYP2U1. ER membrane is a verified part-of descendant and is retained in the specific localization assertions. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: PANTHER:PTN000670347 SUPPORTS TRANSFER Exact current IBD is on the reconstructed human target path; the compartment-granularity interpretation is explained in the reason. |
| GO:0005743 mitochondrial inner membrane | EXP PMID:23176821 Alteration of fatty-acid-metabolizing enzymes affects mitoch... | MODIFY | Summary: Mitochondrial localization is supported; generalize the unresolved inner-membrane assignment to the organelle. Reason: Human GFP-CYP2U1 transiently expressed in HeLa cells partially colocalized with the mitochondrial marker SDHA and the ER marker calregulin (PMID:23176821). This supports mitochondrial association at organelle resolution. The inspected main-text microscopy does not resolve inner versus other mitochondrial membrane locations, and Figure S7 was unavailable. Contact-site and respiration measurements do not supply a direct subcompartment assay. The broader mitochondrion term retains the positive result without treating unresolved inner-membrane precision as evidence against mitochondrial localization. Proposed replacements: mitochondrion |
| GO:0005743 mitochondrial inner membrane | IEA GO_REF:0000044 | MODIFY | Summary: Mitochondrial localization is supported; generalize the unresolved inner-membrane assignment to the organelle. Reason: The literal UniProt SL-0168 assertion on Q7Z449 cites PMID:23176821. That study supports a mitochondrial pool of tagged human CYP2U1 but does not independently resolve inner-membrane residence in the accessible main text. Generalizing to mitochondrion preserves the experimentally supported organelle while keeping the specific membrane boundary and native-protein topology unresolved. Proposed replacements: mitochondrion Supporting Evidence: file:human/CYP2U1/CYP2U1-uniprot.txt file:human/CYP2U1/CYP2U1-source-evidence.yaml |
| GO:0005789 endoplasmic reticulum membrane | EXP PMID:14660610 CYP2U1, a novel human thymus- and brain-specific cytochrome ... | ACCEPT | Summary: CYP2U1 is an ER-associated membrane enzyme. Reason: The original recombinant human enzyme was characterized in heterologous microsomal membranes. Independent human-cell GFP-CYP2U1/calregulin colocalization supports the established ER placement. These observations distinguish heterologous microsomal expression from native human topology and do not establish equivalent localization of both products. |
| GO:0005789 endoplasmic reticulum membrane | IEA GO_REF:0000120 | ACCEPT | Summary: The ER membrane location is independently supported despite a current rule-premise mismatch. Reason: The literal SL-0097 mapping is present in the official record and agrees with human-cell localization. None of the 71 current ARBA00028685 condition branches matches the target feature/taxonomy combination; this does not invalidate the independent location evidence. Supporting Evidence: file:human/CYP2U1/CYP2U1-uniprot.txt file:human/CYP2U1/CYP2U1-source-evidence.yaml |
| GO:0005789 endoplasmic reticulum membrane | TAS Reactome:R-HSA-211960 | ACCEPT | Summary: The specific CYP2U1 arachidonate reaction is assigned to ER membrane. Reason: The exact Reactome 211960 catalyst is CYP2U1 in ER membrane and joins to Q7Z449. This compartment agrees with independently established membrane localization; the event does not resolve membrane topology or the short splice product. |
| GO:0005789 endoplasmic reticulum membrane | TAS Reactome:R-HSA-2161814 | ACCEPT | Summary: The 19-HETE catalyst-set annotation correctly includes ER-associated CYP2U1. Reason: The complete Reactome 2161814 set-member path reaches Q7Z449 in ER membrane. This is a real target membership, corroborated by independent localization rather than transferred merely from other set members. |
| GO:0005789 endoplasmic reticulum membrane | TAS Reactome:R-HSA-2161940 | ACCEPT | Summary: The 20-HETE catalyst-set annotation correctly includes ER-associated CYP2U1. Reason: The complete Reactome 2161940 set-member path reaches the same Q7Z449 membrane entity. The specific modeled reaction remains distinct from those of other enzymes in the set. |
| GO:0005789 endoplasmic reticulum membrane | TAS Reactome:R-HSA-5602242 | ACCEPT | Summary: The gene-level ER membrane placement is retained with disease-variant context explicit. Reason: Reactome 5602242 models defective CYP2U1 variants in this compartment. Independent human-cell localization supports the gene-level ER membrane assertion; the model is not evidence that every pathogenic variant retains normal localization, stability or enzymatic activity. |
| GO:0006805 xenobiotic metabolic process | IBA GO_REF:0000033 | KEEP AS NON CORE | Summary: CYP2U1 can metabolize xenobiotic drugs in a defined recombinant system. Reason: The actual PTN000670347 IBD is on the human path and has experimentally grounded drug-metabolizing P450 descendants. Independent human CYP2U1 nanodisc experiments form sorafenib N-oxide and N-desethyl sunitinib with CPR/NADPH, identified and quantified against standards. Its contribution to drug clearance in vivo and activity of the 168-residue product remain unestablished. Propagation Review Root cause: NO FAILURE NON CORE Sources checked: PANTHER:PTN000670347 SUPPORTS TRANSFER Exact current IBD is on the reconstructed human target path; substrate-specific interpretation is explained in the reason. Supporting Evidence: PMID:41946119 sunitinib is metabolized by both CYP2D6 and CYP2U1. file:human/CYP2U1/CYP2U1-source-evidence.yaml |
| GO:0008202 steroid metabolic process | IBA GO_REF:0000033 | UNDECIDED | Summary: The steroid-process inheritance is phylogenetically genuine, but target steroid chemistry is unresolved. Reason: PTN000670521 is the actual Bilateria ancestor on the human path. The fly phm/Cyp18a1 donors have experimental steroid-biosynthesis or steroid-catabolism annotations, with verified term ancestry. Human CYP2U1 studies establish fatty-acid, lipidated-serotonin and drug oxidation, but the inspected assays contain no steroid-turnover panel. This does not refute steroid activity or establish a lineage-specific loss; the target-specific substrate inference remains uncertain. Propagation Review Root cause: UNRESOLVED Sources checked: PANTHER:PTN000670521 UNRESOLVED Exact current IBD is on the reconstructed human target path; substrate-specific interpretation is explained in the reason. FB:FBgn0004959 SUPPORTS SOURCE BUT NOT TARGET The fly steroid chemistry is experimentally grounded; its retention in human CYP2U1 remains unresolved, not refuted. FB:FBgn0010383 SUPPORTS SOURCE BUT NOT TARGET The fly steroid chemistry is experimentally grounded; its retention in human CYP2U1 remains unresolved, not refuted. |
| GO:0008395 steroid hydroxylase activity | IBA GO_REF:0000033 | UNDECIDED | Summary: The ancestral steroid-hydroxylase assertion cannot yet be resolved for human CYP2U1. Reason: The exact IBD and target descent are valid. Fly experimental evidence includes a steroid-hydroxylase child and a direct Cyp18a1 steroid-hydroxylase annotation. Conserved P450 chemistry does not by itself settle retention of this substrate class in CYP2U1, and no inspected human steroid panel supplies positive or contradictory evidence. Propagation Review Root cause: UNRESOLVED Sources checked: PANTHER:PTN000670521 UNRESOLVED Exact current IBD is on the reconstructed human target path; substrate-specific interpretation is explained in the reason. FB:FBgn0004959 SUPPORTS SOURCE BUT NOT TARGET The fly steroid chemistry is experimentally grounded; its retention in human CYP2U1 remains unresolved, not refuted. FB:FBgn0010383 SUPPORTS SOURCE BUT NOT TARGET The fly steroid chemistry is experimentally grounded; its retention in human CYP2U1 remains unresolved, not refuted. |
| GO:0016705 oxidoreductase activity, acting on paired donors, with incorporation or reduction of molecular oxygen | IEA GO_REF:0000002 | TOO GENERAL | Summary: The paired-donor oxidoreductase parent is correct but does not distinguish the performed reactions. Reason: All four literal InterPro mappings are present and support the heme oxygenase fold. The experimentally resolved fatty-acid and N-arachidonoylserotonin oxygenations provide the more informative catalytic synthesis. |
| GO:0016712 oxidoreductase activity, acting on paired donors, with incorporation or reduction of molecular oxygen, reduced flavin or flavoprotein as one donor, and incorporation of one atom of oxygen | IBA GO_REF:0000033 | TOO GENERAL | Summary: The flavoprotein-supported oxygenase inheritance is sound but broad. Reason: The exact PTN008612736 IBD is on the target path, with experimental descendant grounding including CYP2U1. Human POR-supported reconstitution confirms this donor chemistry. The more specific fatty-acid hydroxylase units and separately proposed lipidated-serotonin activity describe its performed chemistry. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: PANTHER:PTN008612736 SUPPORTS TRANSFER Exact current IBD is on the reconstructed human target path; substrate-specific interpretation is explained in the reason. |
| GO:0016712 oxidoreductase activity, acting on paired donors, with incorporation or reduction of molecular oxygen, reduced flavin or flavoprotein as one donor, and incorporation of one atom of oxygen | IEA GO_REF:0000002 | TOO GENERAL | Summary: The InterPro oxygenase mapping is valid but less specific than the characterized activities. Reason: IPR008069 is on Q7Z449 and its current GO mapping includes this flavoprotein-donor activity. Independent enzyme assays support the general chemistry while resolving distinct substrates and reaction positions. |
| GO:0019369 arachidonate metabolic process | IEA GO_REF:0000041 | ACCEPT | Summary: CYP2U1 directly participates in arachidonate metabolism. Reason: The literal UniPathway UPA00383 cross-reference is present on the official protein record. CYP2U1 itself performs arachidonate 19- and 20-hydroxylation, so this is enzymatic participation rather than a secondary consequence of disease or a substrate-only role. |
| GO:0020037 heme binding | IBA GO_REF:0000033 | ACCEPT | Summary: The inherited heme-binding function agrees with direct CYP2U1 biochemistry. Reason: The exact PTN008612736 IBD is on the verified path, and current donor records contain experimental heme-binding evidence. Recombinant human CYP2U1 is a spectroscopically characterized heme P450. Heme is the catalytic cofactor for the measured oxygenation reactions. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN008612736 SUPPORTS TRANSFER Exact current IBD is on the reconstructed human target path; substrate-specific interpretation is explained in the reason. |
| GO:0020037 heme binding | IEA GO_REF:0000002 | ACCEPT | Summary: The mapped heme-binding sites support the catalytic cofactor function. Reason: All four literal InterPro accessions occur on the official record and map to heme binding. This is independently supported by recombinant heme-dependent P450 chemistry; it does not assign a functional holoenzyme to every alternative product. |
| GO:0052869 obsolete arachidonate omega-hydroxylase activity | EXP PMID:14660610 CYP2U1, a novel human thymus- and brain-specific cytochrome ... | MODIFY | Summary: Replace the obsolete arachidonate-specific term with its current long-chain fatty-acid class. Reason: Current GO marks GO:0052869 obsolete as a substrate-specific duplicate of the broader valid class. The original human 544-residue enzyme converts arachidonate to 20-HETE as well as 19-HETE. The current omega-hydroxylase activity describes the supported terminal chemistry. Proposed replacements: long-chain fatty acid omega-hydroxylase activity |
| GO:0052869 obsolete arachidonate omega-hydroxylase activity | IDA PMID:24563460 Oxidation of endogenous N-arachidonoylserotonin by human cyt... | MODIFY | Summary: Use the current long-chain fatty-acid omega-hydroxylase term for the confirmed arachidonate reaction. Reason: The later human recombinant preparation independently forms the terminal hydroxylation product. GO:0052869 is obsolete, whereas GO:0102033 captures the supported chemistry. Mixed full-length and N-terminally proteolyzed bacterial preparations are not evidence for the natural 168-residue splice product. Proposed replacements: long-chain fatty acid omega-hydroxylase activity |
| GO:0052869 obsolete arachidonate omega-hydroxylase activity | IEA GO_REF:0000116 | MODIFY | Summary: The literal Rhea reaction maps to the current long-chain fatty-acid omega-hydroxylase activity. Reason: RHEA:39755 is explicitly recorded on Q7Z449 for arachidonate conversion to 20-HETE. The reaction is experimentally supported, while its original GO:0052869 identifier is obsolete. The current activity term captures this terminal hydroxylation. Proposed replacements: long-chain fatty acid omega-hydroxylase activity Supporting Evidence: file:human/CYP2U1/CYP2U1-uniprot.txt file:human/CYP2U1/CYP2U1-source-evidence.yaml |
| GO:0097267 omega-hydroxylase P450 pathway | IDA PMID:24563460 Oxidation of endogenous N-arachidonoylserotonin by human cyt... | ACCEPT | Summary: CYP2U1 performs a terminal arachidonate-hydroxylation step in this pathway. Reason: POR/NADPH-supported human enzyme assays identify the 20-HETE product. CYP2U1 therefore catalyzes a step of the existing omega-hydroxylase P450 pathway; no downstream signaling, FAAH regulation or disease process is inferred merely from substrate pharmacology. |
| GO:0097267 omega-hydroxylase P450 pathway | TAS Reactome:R-HSA-2142816 | ACCEPT | Summary: The HETE pathway genuinely contains CYP2U1-catalyzed hydroxylation. Reason: The exact Reactome 2142816 child paths include the CYP2U1-containing 19- and 20-HETE catalyst sets. The terminal 20-HETE step directly supports the existing omega-hydroxylase pathway annotation. Set membership does not assign all 16–20 hydroxylation positions to this protein. Supporting Evidence: Reactome:R-HSA-2142816 file:human/CYP2U1/CYP2U1-source-evidence.yaml |
| GO:0102033 long-chain fatty acid omega-hydroxylase activity | EXP PMID:14660610 CYP2U1, a novel human thymus- and brain-specific cytochrome ... | ACCEPT | Summary: Human CYP2U1 directly catalyzes long-chain fatty-acid omega hydroxylation. Reason: The 544-residue human construct expressed in Sf9 cells with human electron-transfer partners forms LC-MS/MS-identified 20-HETE. DHA oxidation was also observed, but its product positions were unresolved and should not be represented as proven DHA omega regioselectivity. Supporting Evidence: |
| GO:0102033 long-chain fatty acid omega-hydroxylase activity | EXP PMID:24563460 Oxidation of endogenous N-arachidonoylserotonin by human cyt... | ACCEPT | Summary: The later recombinant study independently supports the same omega-hydroxylase chemistry. Reason: Human CYP2U1 reconstitution forms 20-HETE from arachidonate. The bacterial preparation includes N-terminal proteolytic products whose individual activities were not determined; these are distinct from the official short splice product. Supporting Evidence: |
| GO:0102033 long-chain fatty acid omega-hydroxylase activity | IEA GO_REF:0000120 | ACCEPT | Summary: The combined EC/Rhea assignment matches the experimentally established long-chain reaction class. Reason: The current official Q7Z449 catalytic object contains EC 1.14.14.80 and RHEA:56748 for terminal hydroxylation of an omega-methyl long-chain fatty acid. The assignment agrees with human arachidonate turnover and is not inferred solely from a generic P450 family label. |
| GO:0120502 fatty acid omega-1 hydroxylase activity | IEA GO_REF:0000116 | MODIFY | Summary: Refine fatty-acid omega-1 hydroxylation to the established long-chain activity. Reason: RHEA:39759 explicitly converts C20 arachidonate to 19-HETE. GO:0120319 is the valid long-chain omega-1 child of the general source term and matches the measured substrate; no medium-chain substrate range is assumed. Proposed replacements: long-chain fatty acid omega-1 hydroxylase activity |
| GO:1903604 cytochrome metabolic process | TAS Reactome:R-HSA-211958 | MARK AS OVER ANNOTATED | Summary: CYP2U1 catalyzes substrate metabolism, not the cytochrome metabolic process represented by this term. Reason: GO:1903604 describes reactions involving a cytochrome and has protein-metabolic-process ancestry. The literal Reactome 211958 grouping, Miscellaneous substrates, contains CYP enzymes acting on chemically different substrates; its exact CYP2U1 child is arachidonate 19-hydroxylation. That model supports the enzyme acting on fatty acid, not synthesis, modification or degradation of a cytochrome. The over-annotation concerns this process mapping, not the valid underlying reaction. Supporting Evidence: Reactome:R-HSA-211958 Reactome:R-HSA-211960 file:human/CYP2U1/CYP2U1-source-evidence.yaml |
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Download this section (compressed HTML)Q: Which endogenous substrate transformations account for the mitochondrial and neurodevelopmental consequences of CYP2U1 loss?
Q: Does the 168-residue splice product have a stable endogenous protein product and a function distinct from the complete P450?
Q: What is the native submitochondrial location of CYP2U1, and is its ER-associated pool enriched at organelle contact sites?
Q: Does human CYP2U1 retain any steroid-hydroxylating substrate range inferred at the ancestral PAINT node?
Q: Does CYP2U1 catalyze riboflavin oxidation, and what product is formed, beyond the demonstrated binding and modeling observations?
Experiment: In a human neural cell model with measurable endogenous CYP2U1 and substrate, compare wild-type cells, an isogenic CYP2U1 knockout and rescue with the full-length product at matched expression. Quantify endogenous substrate and 2-oxo product by targeted LC-MS/MS using authentic standards and isotope-labeled internal standards, with a low-dose stable-isotope substrate tracing arm to distinguish turnover from altered substrate abundance. Confirm rescue protein abundance and localization. A loss-and-rescue effect on product flux would support cellular participation; a negative result would remain specific to the tested cell state and substrate availability.
Hypothesis: Endogenous human CYP2U1 contributes to cellular conversion of N-arachidonoylserotonin to its 2-oxo product.
Type: Isogenic loss-and-rescue with targeted isotope-tracing lipidomics
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