FA2H is an endoplasmic-reticulum membrane fatty acid 2-hydroxylase that converts free fatty acids to their (R)-2-hydroxy derivatives. The 372-residue protein combines an N-terminal cytochrome-b5 domain with a membrane-associated hydroxylase domain. Its reaction supplies hydroxylated acyl precursors for ceramides, galactosylceramides and glucosylceramides. These lipids support long-term myelin maintenance and context-dependent epidermal membrane formation. Biallelic FA2H variants cause a spectrum of leukodystrophy, hereditary spastic paraplegia 35 and fatty acid hydroxylase-associated neurodegeneration (FAHN), sometimes with brain iron accumulation. A shorter 159-residue isoform lacks the first 213 residues of the longer protein; its activity and localization remain unresolved.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβPEX12 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with O00623 (PEX12). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The contact does not establish that FA2H recruits peroxisomal cargo or provides ubiquitin-ligase activity. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβPGRMC2 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with O15173 (PGRMC2). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The contact does not identify FA2H as a progesterone ligand or establish receptor signaling. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβERLIN1 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with O75477 (ERLIN1). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. An ERLIN1 contact does not establish an ER-associated degradation adaptor function for FA2H. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβSGPL1 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with O95470 (SGPL1). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The interaction does not demonstrate activation of sphingosine-1-phosphate lyase or transfer its catalytic activity to FA2H. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβASGR2 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with P07307-3 (ASGR2). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The receptor contact does not establish a physiological receptor ligand or endocytic function for FA2H. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβCD79A interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with P11912 (CD79A). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The contact does not establish B-cell receptor signaling or a receptor-assembly role for FA2H. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβMSR1 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with P21757 (MSR1). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The scavenger-receptor contact is not an assay of receptor activation, uptake or physiological ligand recognition. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβAQP6 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with Q13520 (AQP6). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The contact does not establish channel regulation or a transport activity of FA2H. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβSLC10A1 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with Q14973 (SLC10A1). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The contact does not establish bile-acid transport or regulation of the transporter by FA2H. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβEBP interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with Q15125 (EBP). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. A sterol-isomerase contact does not demonstrate enzyme activation or a coupled catalytic assembly. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβDLG2 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with Q15700 (DLG2). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The contact does not establish a multipartner adaptor mechanism or synaptic scaffolding by FA2H. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβSLC10A6 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with Q3KNW5 (SLC10A6). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The contact does not establish organic-solute transport or transporter regulation. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβFAM209A interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with Q5JX71 (FAM209A). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. No specific molecular mechanism for this pair is established by the binary screen. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβMARCHF8 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with Q5T0T0 (MARCHF8). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The contact does not establish ubiquitination, ligase regulation or a substrate-adaptor role for FA2H. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβARMC12 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with Q5T9G4-2 (ARMC12). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The contact alone does not establish a functional assembly or trafficking mechanism for FA2H. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβRIC3 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with Q7Z5B4-5 (RIC3). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The contact does not establish acetylcholine-receptor chaperoning by FA2H. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβSYT2 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with Q8N9I0 (SYT2). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The contact does not establish calcium sensing or a synaptic-vesicle adaptor function for FA2H. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβGPR152 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with Q8TDT2 (GPR152). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The receptor contact does not identify FA2H as a signaling ligand or demonstrate receptor activation. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβGPX8 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with Q8TED1 (GPX8). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The contact does not establish peroxidase regulation or transfer GPX8 redox chemistry to FA2H. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβCREB3L1 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with Q96BA8 (CREB3L1). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The screen does not resolve the membrane precursor versus cleaved DNA-binding form of CREB3L1 or establish transcriptional coregulation by FA2H. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβTMEM41A interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with Q96HV5 (TMEM41A). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The binary contact does not establish a membrane-remodeling or adaptor mechanism. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβTLCD4 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with Q96MV1 (TLCD4). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The contact does not establish regulation of lipid transport or a coupled enzymatic role. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβELOVL4 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with Q9GZR5 (ELOVL4). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The contact does not demonstrate an elongase-activator or substrate-channeling function for FA2H. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβTMEM14B interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with Q9NUH8 (TMEM14B). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The binary contact does not establish a transport or membrane-assembly mechanism. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβERGIC3 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with Q9Y282 (ERGIC3). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The contact does not establish cargo recognition or a trafficking-adaptor function for FA2H. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | KEEP AS NON CORE | Summary: HuRI reports a binary FA2HβTMX2 interaction. Reason: PMID:32296183 and its HuRI interaction records explicitly contain FA2H with Q9Y320 (TMX2). The human ORF513/Gal4-fusion experiment was performed in yeast; array, pooling and validated-two-hybrid records describe screen evidence, not three independent native-complex measurements. The contact does not establish oxidoreductase regulation or disulfide chemistry by FA2H. Retain the supported interaction under the campaign exception for non-core generic binding. Historical construct labels do not establish current full-sequence identity or activity of the natural 159-residue product. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32814053 Interactome Mapping Provides a Network of Neurodegenerative ... | KEEP AS NON CORE | Summary: NeuroNet reports FA2H binding to DYNC1H1 in a yeast two-hybrid screen. Reason: PMID:32814053 Table S2, NN1.0 row 3175, records DYNC1H1/FA2H; the current IntAct records identify a FA2H sufficient binding region at residues 93β372. This is neither the displayed full-length protein nor the natural 159-residue product. The pairwise reporter assay supports a contact, but not binding to an assembled dynein motor complex (GO:0070840) or a motor/cargo-adaptor function. Retain generic binding as non-core under the campaign exception; no target-specific DULIP/LuTHy validation or native neuronal complex is claimed. Supporting Evidence: |
| GO:0005515 protein binding | IPI PMID:32814053 Interactome Mapping Provides a Network of Neurodegenerative ... | KEEP AS NON CORE | Summary: NeuroNet reports a FA2HβKLK6 interaction in yeast. Reason: PMID:32814053 Table S2, NN1.0 row 3176, and IntAct explicitly support the KLK6/FA2H pair, with FA2H residues 93β372 recorded as the sufficient binding region. Protease binding (GO:0002020) was considered, but these fusion-screen records do not resolve the KLK6 processing/activity state or establish a functional protease-binding role for FA2H. Retain the observed contact as non-core generic binding under the campaign exception, without inferring protease inhibition, cleavage of FA2H, or equivalence to either current natural product. Supporting Evidence: |
| GO:0005783 endoplasmic reticulum | IBA GO_REF:0000033 | MODIFY | Summary: Conserved ER localization can be refined to the ER membrane. Reason: The actual PTHR12863 tree places the exact 372-residue human target below the asserted PTN000304948 node, with the ER IBD on that path and no intervening NOT. This inherited location is sound. In PMID:15658937, sequence-verified mouse FA2H expressed as an EGFP fusion in CHO cells colocalized with calnexin; the active fusion and membrane-associated protein were examined separately. Human FA2H membrane fractionation in PMID:15337768 and the explicit human ER-membrane catalyst in Reactome support the membrane refinement. This is conserved mouse localization plus human fractionation/model evidence, not human organelle-marker microscopy or localization of the natural 159-residue product. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN000304948 Β· PTN000304948 SUPPORTS TRANSFER The inspected PTHR12863 tree places the exact human 372-residue target below PTN000304948, with the ER IBD and no target-path NOT. The inherited ER location is supported; independent mouse calnexin colocalization, human membrane fractionation and the explicit human Reactome catalyst support refinement to ER membrane. This records target-specific granularity, not erroneous source biology or propagation. Proposed replacements: endoplasmic reticulum membrane |
| GO:0005783 endoplasmic reticulum | IEA GO_REF:0000107 | MODIFY | Summary: The mouse-derived ER annotation supports a more specific ER-membrane location. Reason: The current mouse donor Q5MPP0 has an experimental ER annotation to PMID:15658937. Its original Methods and Results identify sequence-verified mouse constructs in CHO cells, active EGFP-FA2H, and calnexin colocalization. Human membrane fractionation and the explicit human Reactome catalyst provide independent corroboration at ER-membrane resolution. Retain the orthology transfer while refining the compartment; do not recast the mouse imaging as endogenous human evidence or transfer it to the untested short natural product. Proposed replacements: endoplasmic reticulum membrane |
| GO:0005789 endoplasmic reticulum membrane | IEA GO_REF:0000044 | ACCEPT | Summary: The curated location mapping correctly places the characterized FA2H protein in the ER membrane. Reason: The actual SL-0097 record maps directly to GO:0005789. This agrees with human membrane fractionation, conserved mouse EGFP/calnexin localization, and Reactome entity 5693771, which explicitly represents human Q7L5A8 residues 1β372 in the ER membrane. The topology prediction and whole-protein chain feature are not evidence for proteolytic maturation, and the inference does not establish identical localization for the 159-residue product. |
| GO:0005789 endoplasmic reticulum membrane | TAS Reactome:R-HSA-5693761 | ACCEPT | Summary: The actual human reaction catalyst is explicitly an ER-membrane FA2H entity. Reason: The reaction 5693761 links through catalystActivity 5693796 to physicalEntity 5693771: human Q7L5A8 residues 1β372, whose own compartment is ER membrane. This is the target catalyst compartment, not the union of compartments on the parent sphingolipid pathway. Human membrane fractionation and mouse calnexin colocalization are consistent independent evidence. The model does not represent all natural products. |
| GO:0006629 lipid metabolic process | IEA GO_REF:0000002 | MODIFY | Summary: Human FA2H performs fatty-acid metabolism, providing a more informative term than generic lipid metabolism. Reason: IPR014430 genuinely matches the human 372-residue sequence and maps to the imported broad lipid-metabolism term. Selected original Methods and Results in PMIDs:15337768 and 15863841 directly assay hydroxylation of free deuterated tetracosanoic acid by microsomes expressing human FA2H, with GC-MS detection of the hydroxylated product. GO:0006631 is a supported descendant focused on the assayed substrate class. The family includes enzymes with different immediate substrates; neither its Scs7 name nor downstream sphingolipid composition proves human ceramide-bound substrate turnover. Proposed replacements: fatty acid metabolic process |
| GO:0006631 fatty acid metabolic process | IEA GO_REF:0000041 | ACCEPT | Summary: Fatty-acid metabolism is a directly supported substrate-class process for FA2H. Reason: The imported UniPathway UPA00199 link is present in the actual human UniProt record. Independent human free-fatty-acid assays establish that FA2H executes chemical modification of fatty acids, rather than merely being consumed by the pathway. The current term covers that substrate class. This remains distinct from the more specific molecular function and from downstream ceramide and glycosphingolipid biosynthesis. The current standalone UniPathway-to-GO mapping file was not recovered; the literal GOA assertion and target record are preserved without inventing a mapping-file read. |
| GO:0006679 glucosylceramide biosynthetic process | IEA GO_REF:0000107 | ACCEPT | Summary: Conserved FA2H activity contributes hydroxylated acyl precursors to a subset of glucosylceramides. Reason: The actual mouse donor trace reaches the IMP annotation in PMID:21628453. Accessible original Methods and Results distinguish epidermis from whole skin: knockout did not reduce epidermal hydroxyceramides/glucosylceramides at the examined ages, while whole-skin auricle/tail extracts lost or reduced selected 2-hydroxy-acyl glucosylceramides, including C20 and C22β24 species. Retain this bounded biosynthetic role, corroborated by FA2H-dependent hydroxylated HexCer in mouse adipocytes and human free-fatty-acid chemistry. FA2H supplies the hydroxylated acyl precursor; it does not perform glucosyl transfer. This is not evidence for universal dependence of every tissue or molecular species. |
| GO:0006679 glucosylceramide biosynthetic process | ISS GO_REF:0000024 | ACCEPT | Summary: The manually transferred glucosylceramide biosynthetic role is supported with tissue and lipid-species limits. Reason: Current Q5MPP0 donor GOA identifies PMID:21628453 as experimental support. Its selected original Methods/Results show a whole-skin deficit of selected 2-hydroxy glucosylceramides, while isolated epidermis retained the examined species. The ortholog-supported role is production of hydroxylated acyl precursors used in glucosylceramides, not glucosyltransferase chemistry or an assertion that all glucosylceramides require FA2H. Mouse adipocyte lipid profiling in PMID:22517924 and executed human free-fatty-acid hydroxylation provide compatible support. |
| GO:0006682 galactosylceramide biosynthetic process | IEA GO_REF:0000120 | ACCEPT | Summary: FA2H contributes to galactosylceramide biosynthesis by providing hydroxylated acyl precursors. Reason: The combined source explicitly includes rat and mouse orthologs and UPA00787; the human record carries that pathway link. In PMID:17901466, rat Schwann-cell FA2H knockdown reduced fatty-acid 2-hydroxylase activity and hydroxylated GalCer; the activity assay used free deuterated C24 substrate. Mouse FA2H expression in CHO-CGT cells produced hydroxylated HexCer in PMID:15658937, and mouse knockout removed hydroxylated myelin GalCer in PMID:18815260. These performed precursor-supply steps support the process without assigning galactosyltransferase activity, universal GalCer dependence, or immediate ceramide-bound substrate specificity. |
| GO:0006682 galactosylceramide biosynthetic process | ISS GO_REF:0000024 | ACCEPT | Summary: The rat-supported GalCer biosynthetic contribution is retained. Reason: Current rat Q2LAM0 GOA traces this role to IMP evidence in PMID:17901466. Actual primary rat Schwann-cell and D6P2T experiments, free-C24 hydroxylase assays and lipid analyses connect FA2H to hydroxylated GalCer production. The conserved human role is hydroxylated-acyl precursor production; the transfer is not a direct human Schwann-cell experiment and does not make FA2H the glycosyltransferase. Nonhydroxylated GalCer and immediate substrate identity remain separate questions. |
| GO:0006682 galactosylceramide biosynthetic process | ISS GO_REF:0000024 | ACCEPT | Summary: The mouse-supported GalCer biosynthetic contribution is retained. Reason: Mouse Q5MPP0 has experimental GalCer annotations to PMIDs:15658937 and 18815260. Sequence-verified mouse expression in CHO-CGT cells generated hydroxylated HexCer, while knockout lipid analysis demonstrated loss of hydroxylated GalCer with nonhydroxylated compensation. Human free-fatty-acid hydroxylation supplies a coherent conserved biochemical step. These data support biosynthetic precursor supply, not galactose transfer or equivalent activity of both human natural products. |
| GO:0016020 membrane | IDA PMID:15337768 The human FA2H gene encodes a fatty acid 2-hydroxylase. | MODIFY | Summary: Conserved localization evidence refines the membrane-associated enzyme to the ER membrane. Reason: PMID:15337768 Cell Fractionation and Figure 5C directly show recombinant human FA2H in the membrane pellet of COS7 cells; its precise ER location was only predicted. Independent PMID:15658937 Methods and Figure 4 show a sequenced FVB/N mouse 372-residue EGFPβFA2H construct colocalizing with calnexin in CHO-CGT cells, with retained lipid-product formation. Together these support conserved ER-membrane localization (GO:0005789), also represented by the human FA2H entity R-HSA-5693771. The refinement is not a claim of direct human organelle-marker microscopy, measured membrane topology, or localization of the natural 159-residue human product. Proposed replacements: endoplasmic reticulum membrane |
| GO:0016020 membrane | IEA GO_REF:0000002 | MODIFY | Summary: The broad family-derived membrane annotation can be refined to ER membrane. Reason: The actual IPR014430 match spans the human displayed protein and supplies the generic membrane mapping. Human microsomal fractionation, mouse EGFP-FA2H/calnexin colocalization and the target-specific human Reactome ER-membrane entity justify a more precise location. The conserved mouse imaging is identified as such; predicted transmembrane segments and whole-protein chain notation are not experimental topology or cleavage evidence. The natural 159-residue product is not assigned this location by equivalence. Proposed replacements: endoplasmic reticulum membrane |
| GO:0020037 heme binding | IEA GO_REF:0000002 | ACCEPT | Summary: The cytochrome-b5 heme-binding signature supports heme binding in the characterized long protein. Reason: Actual InterPro IPR018506 maps to heme binding and matches the human cytochrome-b5 signature containing the first histidine ligand; UniProt identifies the corresponding N-terminal domain and predicted axial histidines. The human Ξ1β92 construct in PMID:15337768 loses activity, supporting functional importance of this region but not independently measuring heme occupancy. Retain the sequence/domain-based binding inference with that evidential distinction. The 159-residue natural product lacks this domain; no universal product-level heme-binding claim is made. External CYB5B in the Reactome reaction is a separate electron carrier, not a measurement of FA2H-bound heme. |
| GO:0030148 sphingolipid biosynthetic process | TAS Reactome:R-HSA-1660661 | MODIFY | Summary: Refine the supported precursor contribution to glucosylceramide and galactosylceramide biosynthesis. Reason: The Reactome parent pathway includes reaction R-HSA-5693761, where FA2H performs free-fatty-acid hydroxylation. That target reaction establishes precursor supply; it does not itself assay each downstream glycosylceramide branch. Refine the broad sphingolipid process to the two experimentally corroborated branches rather than the shared ceramide parent. Independent branch-specific evidence supports both refinements: mouse FA2H/CHO-CGT and rat Schwann-cell experiments link the precursor reaction to hydroxylated GalCer (PMID:15658937 and PMID:17901466); the human keratinocyte abstract and mouse whole-skin lipid measurements support selected hydroxylated GlcCer species (PMID:17355976 and PMID:21628453). The latter mouse result is distinct from its retained epidermal lipid pool. These are source-resolved contributions through precursor hydroxylation, not FA2H glycosyltransferase activity or equivalent activity of the 159-residue protein. Current typed GO places both replacements below ceramide and sphingolipid biosynthesis, with neither replacement an ancestor of the other. Proposed replacements: glucosylceramide biosynthetic process galactosylceramide biosynthetic process Supporting Evidence: Reactome:R-HSA-1660661 Reactome:R-HSA-5693761 |
| GO:0032286 central nervous system myelin maintenance | IBA GO_REF:0000033 | ACCEPT | Summary: Conserved CNS myelin maintenance is supported by the actual inherited node and mouse loss-of-function evidence. Reason: The exact human target descends from asserted PTN002656510 in the recovered PTHR12863 tree; the matching maintenance IBD is present and no NOT lies on the target path. In PMIDs:18815260 and 21491498, mouse loss of FA2H-dependent hydroxylated myelin lipids permits initially formed myelin but leads to later CNS degeneration; the latter also examines glial conditional deletion. FA2H contributes through enzymatic production of myelin lipid precursors, not by being a passive substrate. Retain the conserved maintenance role without calling this a human perturbation experiment or assuming identical contributions of both human products. Human biallelic-variant studies independently connect FA2H to progressive white-matter disease, SPG35 and FAHN (PMID:19068277, PMID:20104589 and PMID:20853438), corroborating the neural importance of the precursor pathway. Clinical genetics does not itself replace the mouse maintenance experiments or identify the direct molecular step of degeneration; reported mutant catalytic and abundance effects differ. Supporting Evidence: GO_REF:0000033 |
| GO:0044857 plasma membrane raft organization | IEA GO_REF:0000107 | KEEP AS NON CORE | Summary: A positive mouse-adipocyte membrane-raft phenotype is retained as contextual biology. Reason: The Q5MPP0 donor trace reaches PMID:22517924. Actual 3T3-L1 knockdown, cholera-toxin-B FRAP and enantiomer add-back experiments connect FA2H-derived R-2-hydroxy fatty acids to raft mobility and associated lipid composition. This is a specific differentiated adipocyte context, not a demonstration that FA2H is a general raft assembly factor in every cell. Human FA2H expressed in CHO cells produces the R product, supporting conserved chemistry; the downstream membrane-packing mechanism remains proposed. Retain the positive transferred process outside the principal hydroxylase function. |
| GO:0044857 plasma membrane raft organization | ISS GO_REF:0000024 | KEEP AS NON CORE | Summary: The manually transferred membrane-raft organization claim remains supported but contextual. Reason: PMID:22517924 directly examines mouse 3T3-L1 FA2H knockdown, membrane-probe mobility and stereospecific lipid add-back. Its human CHO expression experiment establishes R-2-hydroxy-fatty-acid product formation, not a separate human raft assay. Preserve the ortholog-supported adipocyte role without generalizing the proposed packing mechanism or equating all tissues and natural products. No negative raft conclusion is inferred from distinct mouse myelin detergent-extraction experiments. |
| GO:0046513 ceramide biosynthetic process | IBA GO_REF:0000033 | MODIFY | Summary: Refine the supported precursor contribution to glucosylceramide and galactosylceramide biosynthesis. Reason: The imported ceramide-biosynthesis IBA is valid: the inspected tree places the exact human target below PTN000304948 with its matching IBD and no target-path NOT. Human self-WITH is legitimate descendant evidence. Refine this inherited broad process to both supported glycosylceramide branches; no propagation failure or family-wide substrate restriction is alleged. Independent branch-specific evidence supports both refinements: mouse FA2H/CHO-CGT and rat Schwann-cell experiments link the precursor reaction to hydroxylated GalCer (PMID:15658937 and PMID:17901466); the human keratinocyte abstract and mouse whole-skin lipid measurements support selected hydroxylated GlcCer species (PMID:17355976 and PMID:21628453). The latter mouse result is distinct from its retained epidermal lipid pool. These are source-resolved contributions through precursor hydroxylation, not FA2H glycosyltransferase activity or equivalent activity of the 159-residue protein. Current typed GO places both replacements below ceramide and sphingolipid biosynthesis, with neither replacement an ancestor of the other. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN000304948 Β· PTN000304948 SUPPORTS TRANSFER The inspected target tree/MSA and matching ceramide-biosynthesis IBD support inherited precursor contribution, including legitimate human descendant evidence. Independent GalCer and GlcCer experiments refine target granularity; this is not a failure of ancestral function or source biology. Proposed replacements: glucosylceramide biosynthetic process galactosylceramide biosynthetic process Supporting Evidence: GO_REF:0000033 |
| GO:0046513 ceramide biosynthetic process | IDA PMID:15337768 The human FA2H gene encodes a fatty acid 2-hydroxylase. | MODIFY | Summary: Refine the supported precursor contribution to glucosylceramide and galactosylceramide biosynthesis. Reason: PMID:15337768 Results and Figures 2, 4 and 6 establish human-FA2H-dependent 2-hydroxyceramides and free-fatty-acid hydroxylation in COS7 microsomes. That source supports the broad performed precursor step, while the two specific downstream branches are grounded independently below. Refine the broad ceramide process to both supported branches without attributing both branch assays to the 2004 study. Independent branch-specific evidence supports both refinements: mouse FA2H/CHO-CGT and rat Schwann-cell experiments link the precursor reaction to hydroxylated GalCer (PMID:15658937 and PMID:17901466); the human keratinocyte abstract and mouse whole-skin lipid measurements support selected hydroxylated GlcCer species (PMID:17355976 and PMID:21628453). The latter mouse result is distinct from its retained epidermal lipid pool. These are source-resolved contributions through precursor hydroxylation, not FA2H glycosyltransferase activity or equivalent activity of the 159-residue protein. Current typed GO places both replacements below ceramide and sphingolipid biosynthesis, with neither replacement an ancestor of the other. Proposed replacements: glucosylceramide biosynthetic process galactosylceramide biosynthetic process Supporting Evidence: |
| GO:0046513 ceramide biosynthetic process | IEA GO_REF:0000120 | MODIFY | Summary: Refine the supported precursor contribution to glucosylceramide and galactosylceramide biosynthesis. Reason: The matched ARBA00027419 human FunFam/Eukaryota rule and mouse donor evidence in PMID:22517924 support the broad ceramide precursor contribution. They do not independently establish every branch; specific GalCer and GlcCer support is supplied by the separate sources below. Refine this target annotation without alleging an erroneous rule condition or direct ceramide-bound hydroxylation. Independent branch-specific evidence supports both refinements: mouse FA2H/CHO-CGT and rat Schwann-cell experiments link the precursor reaction to hydroxylated GalCer (PMID:15658937 and PMID:17901466); the human keratinocyte abstract and mouse whole-skin lipid measurements support selected hydroxylated GlcCer species (PMID:17355976 and PMID:21628453). The latter mouse result is distinct from its retained epidermal lipid pool. These are source-resolved contributions through precursor hydroxylation, not FA2H glycosyltransferase activity or equivalent activity of the 159-residue protein. Current typed GO places both replacements below ceramide and sphingolipid biosynthesis, with neither replacement an ancestor of the other. Proposed replacements: glucosylceramide biosynthetic process galactosylceramide biosynthetic process Supporting Evidence: GO_REF:0000120 |
| GO:0061436 establishment of skin barrier | IMP PMID:17355976 Fatty acid 2-hydroxylase, encoded by FA2H, accounts for diff... | KEEP AS NON CORE | Summary: Retain the positive human keratinocyte skin-barrier contribution as a contextual role. Reason: The verified PMID:17355976 abstract explicitly reports human keratinocyte FA2H-siRNA effects: reduced hydroxylase activity and hydroxylated ceramide/GlcCer levels, abnormal lamellar bodies and failure to form normal extracellular lamellar membranes. Together with its reported ordering of free-fatty-acid hydroxylation before ceramide production, this supports a performed lipid-precursor contribution to barrier establishment, not necessity alone. Retain the curatorβs positive IMP assertion as a non-core epidermal context. Operative Methods, knockdown controls and microscopy remain unread; no permeability measurement or complete-paper inspection is inferred. This human experiment is distinct from PMID:21628453 mouse whole-skin/sebaceous findings and its preserved epidermal lipids. Supporting Evidence: |
| GO:0080132 fatty acid 2-hydroxylase activity | IBA GO_REF:0000033 | MODIFY | Summary: The valid inherited hydroxylase activity can be specified as free-fatty-acid 2-hydroxylation. Reason: The exact human sequence lies below the asserted PTN000304948 node with matching hydroxylase IBD and no target-path NOT. The broader family-level term is valid, but the human full-length construct assays in PMIDs:15337768 and 15863841 directly use free deuterated tetracosanoic acid and detect its hydroxylated product by GC-MS. GO:0120520 explicitly specifies free substrate and is a descendant of the imported activity. Retain the inherited function while making this target-supported refinement; do not infer human ceramide-bound substrate activity from other family members or cell lipid composition. Accessible corroboration is the human keratinocyte abstract of PMID:17355976 and the Reactome R-HSA-5693761 reaction summary, which place free-fatty-acid hydroxylation before ceramide synthesis; these do not replace the separately read microsomal substrate assays. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN000304948 Β· PTN000304948 SUPPORTS TRANSFER The inspected PTHR12863 tree places the exact human target below PTN000304948 with the matching hydroxylase IBD and no target-path NOT. Broad fatty acid 2-hydroxylation transfers legitimately, including human descendant evidence. Human free-C24 substrate assays support the more specific free-fatty-acid activity; this does not imply family-wide free-substrate specificity or a bad ancestral assertion. Proposed replacements: free fatty acid 2-hydroxylase activity Supporting Evidence: GO_REF:0000033 Reactome:R-HSA-5693761 |
| GO:0080132 fatty acid 2-hydroxylase activity | IDA PMID:15337768 The human FA2H gene encodes a fatty acid 2-hydroxylase. | MODIFY | Summary: The measured human activity is free-fatty-acid 2-hydroxylation. Reason: PMID:15337768 Experimental Procedures and Figure 6 assay human-FA2H-expressing COS7 microsomes with deuterated free tetracosanoic acid, an NADPH regeneration system and human NADPH:cytochrome P450 reductase. GC/MS identifies the corresponding 2-hydroxy free acid; empty-vector, N-terminal-deletion and electron-supply controls bound attribution. No ATP/CoA is supplied. GO:0120520 therefore describes the tested substrate more precisely than GO:0080132; no natural 159-residue product activity is inferred. Accessible corroboration is the human keratinocyte abstract of PMID:17355976 and the Reactome R-HSA-5693761 reaction summary, which place free-fatty-acid hydroxylation before ceramide synthesis; these do not replace the separately read microsomal substrate assays. Proposed replacements: free fatty acid 2-hydroxylase activity |
| GO:0080132 fatty acid 2-hydroxylase activity | IDA PMID:15863841 A novel method for the measurement of in vitro fatty acid 2-... | MODIFY | Summary: The quantitative assay directly measures hydroxylation of free fatty acid. Reason: PMID:15863841 Methods (Cell Culture; Fatty Acid 2-Hydroxylation Activity Measurement) and Results/Figures 2β7 use microsomes from COS7 cells transfected with the human pcDNA3-hFA2H construct and free deuterated C24 acid. Product retention time and mass fragments establish free 2-hydroxy acid formation, with time/protein/substrate dependence. The paper separately measures endogenous cell and mouse-brain preparations; those are not human-product identification. Refine to GO:0120520, without treating downstream incorporation into ceramide as direct ceramide-substrate turnover. Accessible corroboration is the human keratinocyte abstract of PMID:17355976 and the Reactome R-HSA-5693761 reaction summary, which place free-fatty-acid hydroxylation before ceramide synthesis; these do not replace the separately read microsomal substrate assays. Proposed replacements: free fatty acid 2-hydroxylase activity |
| GO:0080132 fatty acid 2-hydroxylase activity | IDA PMID:22517924 Stereospecificity of fatty acid 2-hydroxylase and differenti... | MODIFY | Summary: Human FA2H produces the R enantiomer of 2-hydroxy fatty acid; independent assays resolve the free substrate. Reason: PMID:22517924 Methods and Figure 1 use stable FLAG-human-FA2H CHO cells and stereochemical lipid analysis to establish R-2-hydroxypalmitate production. The paper does not independently isolate turnover of a ceramide-bound substrate. The specific free-fatty-acid reaction is directly established by the human microsomal assays in PMID:15337768 and PMID:15863841, supporting refinement to GO:0120520. The mouse 3T3-L1 raft, glucose-uptake and lipid-rescue experiments remain separate, and no equivalence of the natural 159-residue product is established. Accessible corroboration is the human keratinocyte abstract of PMID:17355976 and the Reactome R-HSA-5693761 reaction summary, which place free-fatty-acid hydroxylation before ceramide synthesis; these do not replace the separately read microsomal substrate assays. Proposed replacements: free fatty acid 2-hydroxylase activity |
| GO:0080132 fatty acid 2-hydroxylase activity | IEA GO_REF:0000120 | MODIFY | Summary: The combined family/rule/ortholog activity can be refined to the experimentally supported human free substrate. Reason: The actual target matches IPR014430 and the Eukaryota/FunFam condition of ARBA00084402, which annotates broad fatty-acid 2-hydroxylase activity. Mouse and rat donor assays are compatible but are kept distinct from the direct human measurements. Human microsomes expressing the verified construct hydroxylate free C24 substrate in PMIDs:15337768 and 15863841. Use the current specific GO:0120520 term without importing a sphingolipid-bound substrate from the family label, assigning activity to the natural 159-residue product, or treating predicted metal sites as measured occupancy. Accessible corroboration is the human keratinocyte abstract of PMID:17355976 and the Reactome R-HSA-5693761 reaction summary, which place free-fatty-acid hydroxylation before ceramide synthesis; these do not replace the separately read microsomal substrate assays. Proposed replacements: free fatty acid 2-hydroxylase activity Supporting Evidence: GO_REF:0000120 Reactome:R-HSA-5693761 |
| GO:0080132 fatty acid 2-hydroxylase activity | IMP PMID:17355976 Fatty acid 2-hydroxylase, encoded by FA2H, accounts for diff... | MODIFY | Summary: Independent human biochemical evidence supports the specific free-fatty-acid activity. Reason: PMID:17355976 is available here only as a positive abstract reporting FA2H-dependent hydroxylase activity in human keratinocytes; its operative assay and knockdown controls were not inspected. The gene-level activity is independently resolved by actual human-FA2H microsomal free-C24 substrate experiments in PMID:15337768 and PMID:15863841. Refine the broad activity to GO:0120520 on that independent evidence, while preserving the source IMP and its access limit; this does not pretend the unread keratinocyte experiment itself established the substrate. Accessible corroboration is the human keratinocyte abstract of PMID:17355976 and the Reactome R-HSA-5693761 reaction summary, which place free-fatty-acid hydroxylation before ceramide synthesis; these do not replace the separately read microsomal substrate assays. Proposed replacements: free fatty acid 2-hydroxylase activity |
| GO:0080132 fatty acid 2-hydroxylase activity | TAS Reactome:R-HSA-5693761 | MODIFY | Summary: The actual Reactome reaction explicitly supports free-fatty-acid 2-hydroxylase activity. Reason: Reaction 5693761 uses a free 1,2-saturated fatty-acid input and a 2-hydroxy-fatty-acid output, with human FA2H as catalyst and separate cytochrome-b5 redox partners. The current GO:0120520 definition describes that free-substrate reaction more precisely than GO:0080132. Human free-C24 substrate/GC-MS assays independently support the chemistry. This does not grant FA2H the carrierβs redox identity or establish turnover of ceramide-bound acyl chains or the short natural product. Accessible corroboration is the human keratinocyte abstract of PMID:17355976 and the Reactome R-HSA-5693761 reaction summary, which place free-fatty-acid hydroxylation before ceramide synthesis; these do not replace the separately read microsomal substrate assays. Proposed replacements: free fatty acid 2-hydroxylase activity |
| GO:0120520 free fatty acid 2-hydroxylase activity | IDA PMID:15337768 The human FA2H gene encodes a fatty acid 2-hydroxylase. | ACCEPT | Summary: Human FA2H directly hydroxylates a free fatty-acid substrate. Reason: PMID:15337768 Experimental Procedures and Figure 6 identify deuterated free 2-hydroxytetracosanoate formed from free tetracosanoate by human-FA2H-expressing COS7 microsomes. The empty-vector comparison, weak Met93-start preparation and dependence on the added electron-supply system support FA2H attribution. This is a microsomal reconstitution, not a purified isolated-enzyme experiment or a demonstration that both current natural products have the same activity. |
| GO:0120520 free fatty acid 2-hydroxylase activity | IEA GO_REF:0000116 | ACCEPT | Summary: The Rhea-derived free-fatty-acid 2-hydroxylase activity fits the actual human substrate assays. Reason: The captured Rhea records 38551, 38559 and 38855 distinguish palmitate, tetracosanoate and the generalized free-fatty-acid reaction. GO:0120520 explicitly excludes a fatty-acyl chain already within a sphingolipid. Human FA2H microsomal assays use free deuterated C24 substrate with GC-MS product detection, and human CHO expression supports R-product stereochemistry. Retain the specific molecular function; cellular hydroxylated ceramide changes alone are not evidence for the separate ceramide-bound substrate activity. |
| GO:0120521 4-hydroxysphinganine ceramide fatty acyl 2-hydroxylase activity | IDA PMID:22517924 Stereospecificity of fatty acid 2-hydroxylase and differenti... | UNDECIDED | Summary: The cited study does not resolve direct hydroxylation of a ceramide-bound acyl substrate. Reason: GO:0120521 specifies hydroxylation of the fatty acyl chain already attached to 4-hydroxysphinganine. Actual PMID:22517924 Methods/Results establish R-2-hydroxy-fatty-acid production in human-FA2H CHO cells and altered ceramide/hexosylceramide abundance in mouse adipocytes. Those measurements do not distinguish hydroxylation before versus after N-acylation. The directly inspected human assays in PMID:15337768 and PMID:15863841 use free fatty acid. Keep this more specific source assertion undecided: the evidence gap is not proof that FA2H can never use the ceramide substrate. The positive free-substrate ordering in PMID:17355976 and Reactome R-HSA-5693761 strengthens that established route but does not experimentally exclude additional ceramide-bound turnover. Retained UNDECIDED reflects unresolved specific assay attribution, not absence of particular words or a demonstrated citation defect. FlyBase is the literal assigning database for this human IDA row; that provenance alone is not a species error. |
Loading supporting contentβ¦
Download this section (compressed HTML)Q: Which cells produce the natural 159-residue FA2H protein, and how do its stability, localization and activity compare with the 372-residue product independently of the engineered Met93-start construct?
Q: Does human FA2H directly hydroxylate any ceramide-bound acyl substrate, or are the measured hydroxylated sphingolipids supplied entirely through free-fatty-acid hydroxylation and subsequent acyl incorporation under the tested conditions?
Q: Which hydroxylated acyl-chain species and cell-specific precursor pathways account for long-term myelin maintenance and for the different FA2H dependence of whole-skin versus epidermal glucosylceramides?
Q: Which metals occupy the catalytic center of native human FA2H, and how does occupancy relate to activity independently of its cytochrome-b5 heme? The curated two-Zn proposal is similarity-based and should be distinguished from direct human occupancy measurements.
Experiment: Combine isoform-resolved transcript analysis with product-specific targeted proteomics, then compare sequence-verified 372-residue and 159-residue constructs at matched abundance. Measure ER-marker colocalization, membrane association, stability, heme association and isotope-labelled free-fatty-acid product formation with vector and inactive-enzyme controls. Include the Met93-start construct as a separate experimental comparator.
Hypothesis: The natural 159-residue product may differ from the 372-residue protein in abundance, localization or catalytic competence; sequence deletion alone does not determine which properties are retained.
Type: Product-resolved expression, localization and enzyme assays
Experiment: Compare matched isotope-labelled free fatty acids and ceramides in a reconstituted human FA2H membrane system with defined electron supply. Verify substrate presentation, monitor free-acid release, and resolve immediate products by LC-MS before adding downstream acyl-incorporation enzymes. Use inactive FA2H and enzyme-free controls to distinguish hydroxylation from hydrolysis or later lipid synthesis.
Hypothesis: Direct free-fatty-acid turnover can be distinguished from direct ceramide-bound turnover and from downstream incorporation by following the labelled substrate and immediate product in a defined reaction.
Type: Defined substrate-turnover and isotope-tracing assays
Experiment: Trace labelled free fatty acids into defined ceramide, GalCer and GlcCer species after conditional FA2H depletion and rescue in myelinating glial cultures and separately in sebaceous and epidermal models. Compare early lipid flux with later membrane organization and myelin integrity, using measured FA2H activity and matched cell-state controls to distinguish precursor supply from downstream glycosylation or secondary tissue damage.
Hypothesis: Cell-specific supply and incorporation of hydroxylated acyl species may explain why FA2H loss affects selected skin lipids and later myelin integrity differently.
Type: Cell-type-resolved lipid flux and conditional rescue
Experiment: Measure metal stoichiometry and identity in active, sequence-verified human FA2H preparations using orthogonal elemental and spectroscopic methods, with matched purification blanks and activity controls. Test controlled metal depletion and reconstitution while measuring free-fatty-acid product formation and separating cytochrome-b5 heme from the proposed catalytic metal center.
Hypothesis: Direct metal measurements linked to restored activity can distinguish physiological catalytic occupancy from purification-associated metals and similarity-based proposals.
Type: Native metal occupancy and catalytic reconstitution
Loading supporting contentβ¦
Download this section (compressed HTML)Loading supporting contentβ¦
Download this section (compressed HTML)Loading supporting contentβ¦
Download this section (compressed HTML)