ALDH4A1 is the mitochondrial matrix NAD+-dependent delta-1-pyrroline-5-carboxylate dehydrogenase (P5CDH; EC 1.2.1.88). It oxidizes L-glutamate gamma-semialdehyde, the open-chain form in equilibrium with pyrroline-5-carboxylate, to L-glutamate, producing NADH. This is the second and final catalytic step of proline degradation; the same semialdehyde can also arise from ornithine. ALDH4A1 also acts in trans-4-hydroxy-L-proline catabolism by oxidizing the corresponding hydroxylated semialdehyde. Human P5CDH forms a domain-swapped homodimer, with NAD+-binding and catalytic domains and the catalytic cysteine Cys348. Biallelic loss-of-function variants cause type II hyperprolinemia, with proline and pyrroline-5-carboxylate accumulation. A separate noncatalytic activity has been demonstrated in human-cell and purified-protein experiments: ALDH4A1 associates with MPC1-MPC2 and increases reconstituted mitochondrial pyruvate-carrier transport, including with the catalytically inactive S352L variant. MPC1-MPC2 themselves form the translocation pathway and transport without added ALDH4A1; ALDH4A1 is therefore supported as an accessory transport-enhancing factor, while its native occupancy, stoichiometry and contribution across tissues remain unresolved.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0003842 L-glutamate gamma-semialdehyde dehydrogenase activity | IBA GO_REF:0000033 | ACCEPT | Summary: PAINT assigns the conserved P5CDH catalytic activity. Reason: The seeded ancestral node PTN002684348 assigns L-glutamate gamma-semialdehyde dehydrogenase activity. Direct recombinant human kinetics in PMID:22516612 independently establish this defining reaction. The target P30038 in the descendant evidence is legitimate experimental grounding, not circularity; the PAINT tree and node placement were not reconstructed in this audit. Propagation Review Root cause: NO FAILURE CORE Sources checked: MGI:MGI:2443883 UNRESOLVED Mouse descendant evidence recorded in this GOA row's WITH/FROM field. Its individual experimental evidence was not reconstructed separately from the PAINT node; retaining the IBA does not depend on a donor-count argument. PANTHER:PTN002684348 UNRESOLVED Seeded ancestral node retained. Human target activity is directly established and self-evidence is legitimate; the original PAINT tree/MSA and exact node placement were not independently reconstructed. UniProtKB:P30038 SUPPORTS TRANSFER The human target appears among the recorded descendants and has direct experimental P5CDH evidence in PMID:22516612. Its contribution to the ancestral assertion is legitimate grounding, not circular propagation. Supporting Evidence: PMID:22516612 it catalyzes the NAD+-dependent oxidation of L-glutamate-Ξ³-semialdehyde (GSA) to L-glutamate |
| GO:0003842 L-glutamate gamma-semialdehyde dehydrogenase activity | IEA GO_REF:0000120 | ACCEPT | Summary: Combined automated sources identify the established NAD+-dependent P5CDH reaction. Reason: RHEA:30235 and EC:1.2.1.88 match the reaction recorded in human UniProt and measured with recombinant human protein in PMID:22516612. IPR005931 supplies the seeded family mapping. The internal conditions of ARBA00088971 were not recovered; this does not undermine the independently established target reaction. Propagation Review Root cause: NO FAILURE CORE Sources checked: ARBA:ARBA00088971 UNRESOLVED Exact rule ID retained from the combined automated source; internal rule conditions were not recovered. Human catalytic evidence independently establishes the target activity. InterPro:IPR005931 UNRESOLVED Exact seeded InterPro mapping retained. This accession is not listed in the immutable UniProt snapshot and the live entry could not be read; mapping internals remain unresolved, while human P5CDH assays independently support the target judgment. RHEA:30235 SUPPORTS TRANSFER The identical reaction is present in human UniProt: L-glutamate 5-semialdehyde plus NAD+ and water yields glutamate and NADH. Consistent with measured human catalysis. EC:1.2.1.88 SUPPORTS TRANSFER Exact P5CDH catalytic classification in the human UniProt record; independently supported by recombinant human kinetics. Supporting Evidence: PMID:22516612 it catalyzes the NAD+-dependent oxidation of L-glutamate-Ξ³-semialdehyde (GSA) to L-glutamate file:human/ALDH4A1/ALDH4A1-uniprot.txt Reaction=L-glutamate 5-semialdehyde + NAD(+) + H2O = L-glutamate + NADH |
| GO:0005759 mitochondrial matrix | IEA GO_REF:0000044 | ACCEPT | Summary: The subcellular-location mapping places ALDH4A1 in the mitochondrial matrix. Reason: SL-0170 maps the matrix location explicitly recorded in human UniProt. The original human cloning paper describes matrix P5CDH, and the later structural paper uses the same established compartment. Retain this core enzyme location without inferring membrane insertion. Propagation Review Root cause: NO FAILURE CORE Sources checked: UniProtKB-SubCell:SL-0170 SUPPORTS TRANSFER SL-0170 maps the matrix location explicitly recorded in human UniProt and the original human primary abstract. Supporting Evidence: file:human/ALDH4A1/ALDH4A1-uniprot.txt CC -!- SUBCELLULAR LOCATION: Mitochondrion matrix. PMID:8621661 a mitochondrial matrix NAD(+)-dependent dehydrogenase, catalyzes the second step of the proline degradation pathway. |
| GO:0006562 L-proline catabolic process | IEA GO_REF:0000002 | ACCEPT | Summary: The P5CDH family mapping supports proline degradation. Reason: ALDH4A1 itself executes the second catalytic step, oxidizing the semialdehyde generated after proline oxidation. Human cDNA complements P5CDH-deficient yeast and restores growth on proline in PMID:8621661; human kinetics in PMID:22516612 confirm the reaction. This supports the existing process without adding a redundant child term. Propagation Review Root cause: NO FAILURE CORE Sources checked: InterPro:IPR005931 UNRESOLVED Exact seeded InterPro mapping retained. This accession is not listed in the immutable UniProt snapshot and the live entry could not be read; mapping internals remain unresolved, while human P5CDH assays independently support the target judgment. Supporting Evidence: PMID:22516612 it catalyzes the NAD+-dependent oxidation of L-glutamate-Ξ³-semialdehyde (GSA) to L-glutamate PMID:8621661 Both conferred measurable P5CDh activity and the ability to grow on proline as a sole nitrogen source. |
| GO:0016491 oxidoreductase activity | IEA GO_REF:0000002 | MODIFY | Summary: The aldehyde-dehydrogenase family signatures support oxidoreductase activity. Reason: The seeded family signatures correctly identify an oxidoreductase, but the term is less informative than the directly measured human enzyme subtype. Refine to GO:0003842 because PMID:22516612 establishes NAD+-dependent oxidation of L-glutamate gamma-semialdehyde. This preserves the valid redox classification and does not claim the InterPro mapping is biologically false. Propagation Review Root cause: NO FAILURE CORE Sources checked: InterPro:IPR015590 UNRESOLVED Exact seeded InterPro mapping retained. This accession is not listed in the immutable UniProt snapshot and the live entry could not be read; mapping internals remain unresolved, while human P5CDH assays independently support the target judgment. InterPro:IPR016160 SUPPORTS TRANSFER Exact seeded InterPro mapping retained. This catalytic ALDH signature is listed in the immutable human UniProt record; it supports broad oxidoreductase chemistry without alone establishing substrate specificity. InterPro:IPR029510 SUPPORTS TRANSFER Exact seeded InterPro mapping retained. This catalytic ALDH signature is listed in the immutable human UniProt record; it supports broad oxidoreductase chemistry without alone establishing substrate specificity. Proposed replacements: L-glutamate gamma-semialdehyde dehydrogenase activity Supporting Evidence: PMID:22516612 it catalyzes the NAD+-dependent oxidation of L-glutamate-Ξ³-semialdehyde (GSA) to L-glutamate |
| GO:0005515 protein binding | IPI PMID:32814053 Interactome Mapping Provides a Network of Neurodegenerative ... | REMOVE | Summary: The seeded interaction record links ALDH4A1 with RBBP4. Reason: Remove GO:0005515 as an uninformative molecular-function term under the generic-binding policy. The source study performs repeated Y2H screening and pairwise retesting; the immutable UniProt record lists ALDH4A1-RBBP4 with three experiments. The exact supplementary pair result was not independently recovered and no RBBP4-dependent mechanistic ALDH4A1 activity is established here. Removal does not claim that the interaction is false, nor convert it into a scaffold or adaptor function. Supporting Evidence: file:human/ALDH4A1/ALDH4A1-uniprot.txt P30038; Q09028: RBBP4; NbExp=3; |
| GO:0042802 identical protein binding | IPI PMID:22516612 The three-dimensional structural basis of type II hyperproli... | ACCEPT | Summary: Human P5CDH self-associates as a domain-swapped homodimer. Reason: PMID:22516612 measures an approximately 122 kDa human protein by equilibrium analytical ultracentrifugation across three concentrations and three rotor speeds, and identifies the matching dimer in the crystal lattice. The oligomerization domain engages the partner catalytic domain. Retain this specific, experimentally established assembly property of the core enzyme; no separate binding-only core is needed. Supporting Evidence: PMID:22516612 The molar mass is estimated to be 122 kDa (120 β 125 kDa), which is within 2 % of the value predicted for the dimer (124 kDa). |
| GO:0005739 mitochondrion | IDA GO_REF:0000052 | ACCEPT | Summary: HPA immunofluorescence supports mitochondrial localization. Reason: The live HPA subcellular page reports supported mitochondrial localization with HPA006401, including HaCaT and Hep-G2 cells; it also lists a cytosolic pattern in A-431 cells. Retain the seeded mitochondrial assertion at its measured resolution. The independently established matrix localization does not make the broader mitochondrial location non-core or imply that every cell shows identical staining. Supporting Evidence: file:human/ALDH4A1/ALDH4A1-uniprot.txt CC -!- SUBCELLULAR LOCATION: Mitochondrion matrix. |
| GO:0003842 L-glutamate gamma-semialdehyde dehydrogenase activity | IDA PMID:22516612 The three-dimensional structural basis of type II hyperproli... | ACCEPT | Summary: Recombinant human ALDH4A1 directly catalyzes the P5CDH reaction. Reason: PMID:22516612 measures NADH production with recombinant human P5CDH and P5C/NAD+ substrates, including kinetic analysis and activity loss in S352L. Ligand-complex structures in that study use mouse P5CDH, while the human enzyme kinetics and human apo/mutant structures directly support this IDA. Retain the defining catalytic function. Supporting Evidence: PMID:22516612 it catalyzes the NAD+-dependent oxidation of L-glutamate-Ξ³-semialdehyde (GSA) to L-glutamate PMID:22516612 P5CDH activity was measured at 20 Β°C by monitoring the production of NADH at 340 nm. |
| GO:0005739 mitochondrion | HTP PMID:34800366 Quantitative high-confidence human mitochondrial proteome an... | ACCEPT | Summary: Human mitochondrial proteomics supports the enzyme compartment. Reason: PMID:34800366 uses subtractive/spatial proteomics and mitochondrial importomics; its full main text is available. The exact ALDH4A1 supplementary row was not separately extracted here. Retain the curator-selected HTP mitochondrial annotation with explicit deference, corroborated by independent human P5CDH and HPA localization evidence; no matrix-level precision is attributed to this HTP row. Supporting Evidence: file:human/ALDH4A1/ALDH4A1-uniprot.txt CC -!- SUBCELLULAR LOCATION: Mitochondrion matrix. PMID:34800366 For subtractive proteomics, we compared crude and pure mitochondrial fractions from HEK293T cells by quantitative MS |
| GO:0004029 aldehyde dehydrogenase (NAD+) activity | IDA PMID:4015840 Biochemical genetic analysis of human and rodent aldehyde de... | KEEP AS NON CORE | Summary: Retain the curated broad aldehyde-dehydrogenase assay as a secondary substrate capacity. Reason: The original CACAO8806 entry for human AL4A1/P30038 traces this assertion to Table 1 agar-overlay assays using propionaldehyde or benzaldehyde (https://gowiki.tamu.edu/wiki/index.php/PMID%3A4015840). That assay scope fits the broad NAD+-dependent aldehyde-dehydrogenase term. Retain the experimental curator's source assignment as a secondary substrate capacity, without claiming physiological turnover of those substrates. The primary table remains unread, so exact isozyme attribution is curator-deferred; the abstract's emphasis on other ALDH forms is not evidence of misattribution. Independent GSA kinetics in PMID:22516612 establish the separately annotated catalytic core, but do not justify replacing this assay's substrate scope with GSA-specific chemistry. Supporting Evidence: |
| GO:0005759 mitochondrial matrix | TAS Reactome:R-HSA-6784402 | ACCEPT | Summary: Reactome assigns matrix ALDH4A1 to a hydroxyproline-catabolic reaction. Reason: The preserved Reactome assertion places the catalyst in the matrix; its cached reaction summary explicitly identifies mitochondrial ALDH4A1 and the corresponding NAD+-dependent oxidation. The summary alone does not resolve submitochondrial topology, but the original human P5CDH paper and UniProt independently establish matrix localization. Accept the source-level compartment without treating the three reaction records as separate localization experiments. Supporting Evidence: Reactome:R-HSA-6784402 Mitochondrial delta-1-pyrroline-5-carboxylate dehydrogenase (ALDH4A1) catalyzes the reaction sequence PMID:8621661 a mitochondrial matrix NAD(+)-dependent dehydrogenase, catalyzes the second step of the proline degradation pathway. |
| GO:0005759 mitochondrial matrix | TAS Reactome:R-HSA-70679 | ACCEPT | Summary: Reactome assigns matrix ALDH4A1 to a proline-catabolic reaction. Reason: The preserved Reactome assertion places the catalyst in the matrix; its cached reaction summary explicitly identifies mitochondrial ALDH4A1 and the corresponding NAD+-dependent oxidation. The summary alone does not resolve submitochondrial topology, but the original human P5CDH paper and UniProt independently establish matrix localization. Accept the source-level compartment without treating the three reaction records as separate localization experiments. Supporting Evidence: Reactome:R-HSA-70679 Mitochondrial delta-1-pyrroline-5-carboxylate dehydrogenase (ALDH4A1) catalyzes the reaction of L-glutamate gamma-semialdehyde and NAD+ PMID:8621661 a mitochondrial matrix NAD(+)-dependent dehydrogenase, catalyzes the second step of the proline degradation pathway. |
| GO:0005759 mitochondrial matrix | TAS Reactome:R-HSA-9929439 | ACCEPT | Summary: Reactome assigns matrix ALDH4A1 to a hydroxyproline-catabolic reaction. Reason: The preserved Reactome assertion places the catalyst in the matrix; its cached reaction summary explicitly identifies mitochondrial ALDH4A1 and the corresponding NAD+-dependent oxidation. The summary alone does not resolve submitochondrial topology, but the original human P5CDH paper and UniProt independently establish matrix localization. Accept the source-level compartment without treating the three reaction records as separate localization experiments. Supporting Evidence: Reactome:R-HSA-9929439 Mitochondrial ALDH4A1 catalyzes the reaction of 1PYR-3OH-5COOH PMID:8621661 a mitochondrial matrix NAD(+)-dependent dehydrogenase, catalyzes the second step of the proline degradation pathway. |
| GO:0019470 trans-4-hydroxy-L-proline catabolic process | TAS PMID:21998747 Structural and biochemical studies of human 4-hydroxy-2-oxog... | ACCEPT | Summary: ALDH4A1 executes the second step of trans-4-hydroxy-L-proline catabolism. Reason: The cited HOGA1 paper explicitly places P5CDH second in the four-enzyme mitochondrial pathway; its own assays characterize HOGA1. PMID:22516612 likewise describes the established P5CDH hydroxy-substrate reaction and models hydroxyl accommodation using mouse ligand structures, rather than directly measuring human OH-GSA turnover. Retain the TAS pathway assignment, supported by the consistent curated Reactome hydroxyproline reactions, with this distinction between pathway attribution and newly performed assays. Supporting Evidence: PMID:21998747 hydroxyproline oxidase (HPOX), Ξ1-pyrroline-5-carboxylate dehydrogenase (1P5CDH), aspartate aminotransferase (AspAT), and 4-hydroxy-2-oxoglutarate aldolase (HOGA PMID:22516612 P5CDH is also the second enzyme of hydroxyproline catabolism in humans |
| GO:0003842 L-glutamate gamma-semialdehyde dehydrogenase activity | TAS PMID:8621661 Cloning, characterization, and expression of cDNAs encoding ... | ACCEPT | Summary: Human cDNA complementation establishes P5CDH catalytic identity. Reason: PMID:8621661 reports two human cDNAs encoding the same 563-residue protein; both confer measurable P5CDH activity and proline-dependent growth in a deficient yeast strain. This is direct human-gene functional evidence in a heterologous host, independently supported by later purified-human kinetics. Preserve the seeded TAS code. Supporting Evidence: PMID:8621661 Both conferred measurable P5CDh activity and the ability to grow on proline as a sole nitrogen source. |
| GO:0005759 mitochondrial matrix | TAS PMID:8621661 Cloning, characterization, and expression of cDNAs encoding ... | ACCEPT | Summary: The original human P5CDH paper places the enzyme in the mitochondrial matrix. Reason: The accessible primary abstract explicitly describes a mitochondrial matrix NAD+-dependent dehydrogenase. Preserve the TAS location, independently consistent with curated UniProt and subsequent structural work; the abstract is not presented as a newly inspected import or fractionation assay. Supporting Evidence: PMID:8621661 a mitochondrial matrix NAD(+)-dependent dehydrogenase, catalyzes the second step of the proline degradation pathway. file:human/ALDH4A1/ALDH4A1-uniprot.txt CC -!- SUBCELLULAR LOCATION: Mitochondrion matrix. |
| GO:0141109 transporter activator activity | IDA PMID:40355545 ALDH4A1 functions as an active component of the MPC complex ... | NEW | Summary: ALDH4A1 binds MPC1-MPC2 and increases its reconstituted pyruvate transport. Reason: The live GO:0141109 definition requires binding to and increasing transporter activity. PMID:40355545 provides recombinant-protein binding/complex assays plus Figure 7 proteoliposomes comparing MPC1-MPC2 alone with addition of wild-type human ALDH4A1 or inactive S352L; both increase radiolabel uptake over time. Human-cell association-defective deletion and catalytic-mutant rescue experiments corroborate a noncatalytic contribution. ALDH4A1 itself supplies the interacting accessory protein, rather than merely being a metabolic substrate. Propose only this bounded MF: no standalone pyruvate-transporter activity, fixed-stoichiometry complex or additional transport-process annotation. MPC amount/orientation normalization and the precise distinction between stabilization during reconstitution and kinetic activation remain limitations; other matrix-protein controls and native stoichiometry would strengthen the mechanism. This reported moonlighting activity is kept outside the defining catalytic core. Supporting Evidence: PMID:40355545 Remarkably, wild-type ALDH4A1 or ALDH4A1(S352L) could markedly enhance pyruvate transport into proteoliposomes by MPC1βMPC2 PMID:40355545 In vitro binding assay demonstrated the direct interaction of ALDH4A1 with both MPC1 and MPC2 |
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Download this section (compressed HTML)Q: What fraction of native mitochondrial MPC1-MPC2 carries ALDH4A1, with what stoichiometry and ALDH4A1 oligomeric state, and how does this vary across tissues?
Q: Does ALDH4A1 increase MPC1-MPC2 transport turnover at fixed incorporated transporter amount/orientation, stabilize the complex during reconstitution, or both?
Q: Does the interaction-defective ALDH4A1 deletion preserve purified P5CDH kinetics and native dimer assembly, beyond retaining the reported cellular proline-regulation phenotype?
Experiment: Compare MPC1-MPC2 proteoliposomes with WT ALDH4A1, S352L, a binding-site mutant and an unrelated matrix protein. Quantify incorporated MPC1/MPC2, orientation, vesicle integrity and initial transport rates. Pair this with native-mitochondrial occupancy/stoichiometry measurements and independent validation of mutant catalytic activity and dimerization.
Hypothesis: ALDH4A1 increases transport by binding and stabilizing or activating the MPC heterodimer.
Type: quantitative transport reconstitution and native complex analysis
What is not known β curated, literature-grounded statements of the open unknowns (the inverse of core functions).
Gap: ALDH4A1 binding and enhancement of MPC1-MPC2 transport are supported experimentally, but the native occupancy, stoichiometry and precise activation mechanism remain unresolved.
OPEN BIOLOGY MF_DARK
What is known: Human enzyme catalysis/dimerization and the two-subunit MPC transport pathway are established. PMID:40355545 adds biochemical and human-cell evidence for an accessory contribution independent of catalytic S352L activity. Its gel filtration/BN-PAGE association data do not determine a unique native stoichiometry, and the proteoliposome enhancement does not distinguish altered carrier incorporation/stability from increased turnover of an equal amount of correctly oriented carrier.
Significance: Defines how the catalytic enzyme and accessory transport-enhancing roles coexist and how broadly the second role operates across tissues.
What would resolve it: Quantify native ALDH4A1:MPC occupancy and oligomeric state; normalize reconstituted transport to incorporated MPC amount and orientation and compare with unrelated matrix-protein controls and specific binding-site mutants.
Provenance (the field's own admissions):
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