AASDHPPT is a monomeric, magnesium-dependent 4'-phosphopantetheinyl transferase that uses coenzyme A to install 4'-phosphopantetheine on conserved serines in a broad set of carrier-protein and dehydrogenase acceptors. Its directly established human cytosolic pool matures the ACP domain of FASN and ALDH1L1, while purified human enzyme also modifies mitochondrial acceptors including NDUFAB1 and ALDH1L2. Experiments on the mouse ortholog in C2C12 myoblasts support an additional mitochondrial-matrix pool that phosphopantetheinylates NDUFAB1 and enables mitochondrial fatty acid synthesis and oxidative metabolism; this compartmental role has not yet been demonstrated directly in human cells. The catalogued truncated isoform 2 lacks much of the catalytic region, and its protein abundance, activity, and localization remain unknown.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0008897 holo-[acyl-carrier-protein] synthase activity | IBA GO_REF:0000033 | ACCEPT | Summary: AASDHPPT is the human broad-specificity 4'-phosphopantetheinyl transferase, and holo-[acyl-carrier-protein] synthase activity accurately captures its core catalytic reaction. Reason: The PAINT inference is consistent with the conserved phosphopantetheinyl transferase family and is independently established for the human protein by biochemical and structural studies. The term describes transfer of the phosphopantetheine group from CoA to a carrier-protein serine and is at the right level of specificity for AASDHPPT's core molecular function. Propagation Review Root cause: NO FAILURE CORE Sources checked: CGD:CAL0000182911 SUPPORTS TRANSFER Curated fungal phosphopantetheinyl-transferase family member. PomBase:SPAC17C9.02c SUPPORTS TRANSFER Curated fission-yeast orthologue with the conserved transferase role. SGD:S000003122 SUPPORTS TRANSFER Curated budding-yeast LYS5 orthologue. UniProtKB:G5EB87 SUPPORTS TRANSFER Experimentally characterized fungal phosphopantetheinyl transferase. UniProtKB:P37623 SUPPORTS TRANSFER Bacterial AcpT phosphopantetheinyl transferase. UniProtKB:P39135 SUPPORTS TRANSFER Bacterial Sfp phosphopantetheinyl transferase. UniProtKB:Q4X1W0 SUPPORTS TRANSFER Fungal phosphopantetheinyl transferase in the same conserved family. dictyBase:DDB_G0285927 SUPPORTS TRANSFER Curated Dictyostelium family member. PANTHER:PTN000259224 NOT RELEVANT PAINT ancestral node rather than an independent experimental donor. UniProtKB:Q9NRN7 SUPPORTS TRANSFER Self-reference: the target is its own IBD seed, which is expected rather than circular -- its own IDA/TAS annotation to this term is one of the descendant evidences behind the IBD. The IBA then asserts the additional claim that the function is inherited rather than lineage-specific. Supporting Evidence: PMID:18022563 This enzyme exhibits a broad substrate specificity and is able to phosphopantetheinylate the ACP components of the cytosolic and the mitochondrial FAS systems, as well as aminoadipate semialdehyde dehydrogenase, the human ortholog to the yeast lys5 gene [2, 9]. |
| GO:0051604 protein maturation | IBA GO_REF:0000033 | MODIFY | Summary: The inferred process is biologically sound but protein maturation is much broader than the demonstrated covalent phosphopantetheinylation reaction. Reason: AASDHPPT matures apo-carrier proteins by transferring 4'-phosphopantetheine specifically to a peptidyl-serine. GO:0018070 peptidyl-serine phosphopantetheinylation captures that mechanism directly, whereas GO:0051604 groups many unrelated routes to protein functional maturation. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: PANTHER:PTN000259224 SUPPORTS SOURCE BUT NOT TARGET The ancestral node supports a protein-maturation role, but the human reaction is known precisely enough to use phosphopantetheinylation. UniProtKB:Q9NRN7 SUPPORTS TRANSFER Self-reference: the target is its own IBD seed, which is expected rather than circular -- its own IDA annotation to this term is one of the descendant evidences behind the IBD. The IBA then asserts the additional claim that the function is inherited rather than lineage-specific. Proposed replacements: peptidyl-serine phosphopantetheinylation Supporting Evidence: PMID:19933275 Phosphopantetheinylation by PPT occurs specifically at serine 354 of FDH because replacement of this serine with alanine prevented this covalent modification. |
| GO:0005829 cytosol | IBA GO_REF:0000033 | ACCEPT | Summary: Cytosol is a directly supported active compartment for AASDHPPT and is compatible with modification of both cytosolic and pre-import substrates. Reason: Biochemical fractionation localized the human enzyme to the cytosolic compartment, and the structural study explicitly discusses cytosolic AASDHPPT acting on carrier proteins that may subsequently enter mitochondria. The annotation does not assert that cytosol is the only possible compartment. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN000259224 NOT RELEVANT PAINT ancestral node; human direct evidence independently supports the term. UniProtKB:Q9NRN7 SUPPORTS TRANSFER Self-reference: the target is its own IBD seed, which is expected rather than circular -- its own IDA/TAS annotation to this term is one of the descendant evidences behind the IBD. The IBA then asserts the additional claim that the function is inherited rather than lineage-specific. Supporting Evidence: PMID:18022563 Subcellular fractionation studies indicate that the human PPT is located in the cytosolic compartment, implying that the mitochondrial ACP and aminoadipate semialdehyde dehydrogenase undergo posttranslational modification prior to their import into mitochondria. |
| GO:0000287 magnesium ion binding | IEA GO_REF:0000002 | ACCEPT | Summary: Magnesium binding is a genuine catalytic cofactor property of human AASDHPPT. Reason: The InterPro-based inference is corroborated by direct structural and kinetic characterization: Mg2+ is bound with CoA in the active site and is required for catalysis. This is a core enabling molecular function rather than a nonspecific ion association. Propagation Review Root cause: NO FAILURE CORE Sources checked: InterPro:IPR008278 SUPPORTS TRANSFER Conserved phosphopantetheinyl-transferase domain mapping. InterPro:IPR037143 SUPPORTS TRANSFER Structural/domain family mapping consistent with the human Mg2+-bound structure. Supporting Evidence: PMID:18022563 Molecular cloning and biochemical characterization of human PPT revealed that this cytosolic enzyme is a Mg2+-requiring monomeric enzyme that exhibits a broad acceptor substrate specificity [9]. |
| GO:0005829 cytosol | IEA GO_REF:0000044 | ACCEPT | Summary: The UniProt subcellular-location mapping to cytosol is correct and agrees with direct fractionation evidence. Reason: This IEA is a faithful translation of the reviewed UniProt cytosol record, and primary evidence places human AASDHPPT in the cytosolic compartment. It does not exclude additional pools reported under other conditions. Propagation Review Root cause: NO FAILURE CORE Sources checked: UniProtKB-SubCell:SL-0091 SUPPORTS TRANSFER Direct mapping of the reviewed UniProt cytosol vocabulary term. Supporting Evidence: PMID:18022563 Subcellular fractionation studies indicate that the human PPT is located in the cytosolic compartment, implying that the mitochondrial ACP and aminoadipate semialdehyde dehydrogenase undergo posttranslational modification prior to their import into mitochondria. |
| GO:0008897 holo-[acyl-carrier-protein] synthase activity | IEA GO_REF:0000120 | ACCEPT | Summary: The combined electronic annotation correctly identifies AASDHPPT's core holo-[acyl-carrier-protein] synthase activity. Reason: Independent family, reaction, and EC mappings converge on the same phosphopantetheinyl-transferase reaction, and purified human AASDHPPT has been directly characterized with CoA and ACP. There is no indication of a paralog or pseudoenzyme transfer error. Propagation Review Root cause: NO FAILURE CORE Sources checked: ARBA:ARBA00098018 SUPPORTS TRANSFER Automated rule agrees with direct human enzymology. InterPro:IPR008278 SUPPORTS TRANSFER Conserved phosphopantetheinyl-transferase domain mapping. InterPro:IPR037143 SUPPORTS TRANSFER Structural/domain family mapping supports catalytic conservation. RHEA:12068 SUPPORTS TRANSFER Curated reaction corresponding to the experimentally demonstrated transfer. EC:2.7.8.7 SUPPORTS TRANSFER Enzyme classification matches the reviewed UniProt catalytic activity. Supporting Evidence: PMID:18022563 This enzyme exhibits a broad substrate specificity and is able to phosphopantetheinylate the ACP components of the cytosolic and the mitochondrial FAS systems, as well as aminoadipate semialdehyde dehydrogenase, the human ortholog to the yeast lys5 gene [2, 9]. |
| GO:0005515 protein binding | IPI PMID:16189514 Towards a proteome-scale map of the human protein-protein in... | MARK AS OVER ANNOTATED | Summary: A high-throughput binary screen reported an interaction with TRAF2, but generic protein binding does not describe AASDHPPT's molecular function. Reason: The IntAct-derived pair should be retained as interaction provenance, but an interaction detected in a proteome-scale yeast two-hybrid screen does not establish a stable complex, regulatory mechanism, or binding-specific activity for AASDHPPT. Its informative molecular function is the phosphopantetheinyl-transferase reaction. Supporting Evidence: PMID:16189514 detected approximately 2,800 interactions. |
| GO:0005515 protein binding | IPI PMID:19615732 Defining the human deubiquitinating enzyme interaction lands... | MARK AS OVER ANNOTATED | Summary: USP22-associated proteomics supplies an interaction observation, not an informative binding function for AASDHPPT. Reason: The original study generated a broad candidate DUB-interaction landscape. The AASDHPPT-USP22 association can remain in the interaction record, but absent a demonstrated mechanistic consequence it should not be promoted to a core or specific molecular-function annotation. Supporting Evidence: PMID:19615732 proteins associated with 75 Dubs. |
| GO:0005515 protein binding | IPI PMID:25416956 A proteome-scale map of the human interactome network. | MARK AS OVER ANNOTATED | Summary: The normalized tuple represents high-throughput binary interactions with TRAF2, SIAH1, and USP22; these do not define a generic binding function. Reason: The underlying yeast two-hybrid map supports physical-interaction records, but it does not show that these partners are physiological substrates, regulators, or stable complex members of AASDHPPT. GO:0005515 is therefore uninformative and should not be treated as a core molecular function. Supporting Evidence: PMID:25416956 binary protein-protein interactions. |
| GO:0005515 protein binding | IPI PMID:28514442 Architecture of the human interactome defines protein commun... | MARK AS OVER ANNOTATED | Summary: BioPlex reported a USP22 isoform-2 co-association, but the generic protein binding term adds no mechanistic information. Reason: Affinity-purification mass spectrometry identifies candidate co-associations and does not by itself establish direct binding or a stable AASDHPPT-containing complex. The pair is useful interaction provenance, whereas GO:0005515 should not represent the enzyme's core function. Supporting Evidence: PMID:28514442 With more than 56,000 candidate interactions, |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | MARK AS OVER ANNOTATED | Summary: HuRI reports binary interactions with CRX, TRAF2, EFHC2, and USP22 isoform 2, but these screening hits do not define AASDHPPT's function. Reason: The source validates a reference binary-interaction map, while explicitly leaving the cellular function of most individual pairs unresolved. The exact WITH/FROM partners should be preserved, but no substrate, regulatory, or stable-complex role has been shown for these pairs, making generic protein binding an over-annotation. Supporting Evidence: PMID:32296183 the cellular function of most individual PPIs remains to be elucidated. |
| GO:0005515 protein binding | IPI PMID:33961781 Dual proteome-scale networks reveal cell-specific remodeling... | MARK AS OVER ANNOTATED | Summary: Cell-line interactome proteomics reports association with USP22, but not a specific AASDHPPT binding activity or mechanism. Reason: BioPlex 3.0 is a large affinity-purification interaction network whose associations can vary by cellular context. Without targeted validation of a functional AASDHPPT-USP22 relationship, the interaction should not be elevated to the generic molecular function protein binding. Supporting Evidence: PMID:33961781 interactions among 14,586 proteins. |
| GO:0015939 pantothenate metabolic process | TAS Reactome:R-HSA-199220 | MODIFY | Summary: Placement of AASDHPPT in Reactome's vitamin B5 pathway reflects use of CoA-derived phosphopantetheine, but pantothenate metabolism is not the reaction catalyzed by this enzyme. Reason: Pantothenate is an upstream precursor of CoA. AASDHPPT neither transforms pantothenate nor controls its synthesis; it transfers the phosphopantetheine moiety of CoA to a serine in protein substrates. The mechanistically accurate biological-process term is GO:0018070 peptidyl-serine phosphopantetheinylation. Proposed replacements: peptidyl-serine phosphopantetheinylation Supporting Evidence: Reactome:R-HSA-199220 Vitamin B5 ((R)-pantothenate, PanK), is an essential precursor for the synthesis of the metabolic cofactor Coenzyme A (CoA-SH) (Robishaw and Neely 1985) and is the prosthetic group of acyl carrier protein (ACP) (Joshi et al. 2003). PMID:19933275 Phosphopantetheinylation by PPT occurs specifically at serine 354 of FDH because replacement of this serine with alanine prevented this covalent modification. |
| GO:0008897 holo-[acyl-carrier-protein] synthase activity | TAS Reactome:R-HSA-199202 | ACCEPT | Summary: Reactome explicitly assigns AASDHPPT the transfer of phosphopantetheine from CoA to the ACP domain of FASN, matching the GO molecular function. Reason: The curated reaction directly describes holo-ACP formation and is consistent with biochemical and structural evidence for the human enzyme. This is a core catalytic function rather than merely pathway membership. Supporting Evidence: Reactome:R-HSA-199202 Cytosolic AASDHPPT (alpha-aminoadipic semialdehyde dehydrogenase-phosphopantetheinyl transferase) catalyzes the transfer of a phosphopantetheine moiety from coenzyme A to serine 2156 within the ACP domain of FAS (fatty acyl synthase). |
| GO:0008897 holo-[acyl-carrier-protein] synthase activity | IDA PMID:19933275 Acyl carrier protein-specific 4'-phosphopantetheinyl transfe... | ACCEPT | Summary: Purified human AASDHPPT converts apo-ALDH1L1/FDH to the active holoenzyme, directly supporting phosphopantetheinyl-transferase activity. Reason: Recombinant-enzyme and cellular knockdown experiments identify AASDHPPT as the enzyme that transfers 4'-phosphopantetheine to the carrier-protein-like domain of FDH. Although FDH is not a canonical stand-alone ACP, the GO term definition encompasses ACP-like carrier-protein substrates and accurately captures the demonstrated reaction. Supporting Evidence: PMID:19933275 In the current study, we demonstrate that the broad specificity human PPT converts apo-FDH to holoenzyme and thus activates FDH catalysis. |
| GO:0008897 holo-[acyl-carrier-protein] synthase activity | IDA PMID:21238436 Enzymatic properties of ALDH1L2, a mitochondrial 10-formylte... | ACCEPT | Summary: Recombinant AASDHPPT phosphopantetheinylates mitochondrial ALDH1L2/mtFDH at a defined acceptor serine. Reason: Direct reactivation and site-directed mutagenesis show transfer to Ser375 of the ACP-like intermediate domain of ALDH1L2. This independent substrate study reinforces the broad carrier-protein phosphopantetheinyl-transferase activity of AASDHPPT. Supporting Evidence: PMID:21238436 Our results, however, demonstrate that PPT modifies mtFDH exclusively at Ser375. |
| GO:0009258 10-formyltetrahydrofolate catabolic process | IMP PMID:19933275 Acyl carrier protein-specific 4'-phosphopantetheinyl transfe... | KEEP AS NON CORE | Summary: AASDHPPT phosphopantetheinylation activates ALDH1L1/FDH and thereby acts upstream of its 10-formyltetrahydrofolate catabolic reaction. Reason: The directional qualifier is important: AASDHPPT does not itself catabolize 10-formyltetrahydrofolate, but its modification of FDH restores that enzyme's dehydrogenase activity. This is a valid substrate-specific downstream consequence of the core transferase function, not the core reaction of AASDHPPT itself. Supporting Evidence: PMID:19933275 In our experiments, incubation of this enzyme with purified recombinant human PPT in the presence of CoA and Mg 2+ restored 10-fTHF dehydrogenase activity. |
| GO:0009258 10-formyltetrahydrofolate catabolic process | IMP PMID:21238436 Enzymatic properties of ALDH1L2, a mitochondrial 10-formylte... | KEEP AS NON CORE | Summary: AASDHPPT modification of ALDH1L2/mtFDH enables mitochondrial 10-fTHF dehydrogenase catalysis. Reason: AASDHPPT acts upstream by converting ALDH1L2 to its active phosphopantetheinylated form; the substrate enzyme, not AASDHPPT, performs the folate-catabolic chemistry. The annotation is therefore biologically valid with its causal qualifier but represents a secondary consequence of the transferase reaction. Supporting Evidence: PMID:21238436 As in the case of the cytosolic enzyme, this modification enables 10-fTHF dehydrogenase catalysis of mtFDH. |
| GO:0051604 protein maturation | IDA PMID:19933275 Acyl carrier protein-specific 4'-phosphopantetheinyl transfe... | MODIFY | Summary: AASDHPPT does mature apo-FDH to holo-FDH, but protein maturation is too broad for the experimentally defined modification. Reason: The paper identifies covalent addition of 4'-phosphopantetheine at FDH Ser354. GO:0018070 names this peptidyl-serine modification directly and avoids conflating it with unrelated mechanisms covered by the parent term protein maturation. Proposed replacements: peptidyl-serine phosphopantetheinylation Supporting Evidence: PMID:19933275 Phosphopantetheinylation by PPT occurs specifically at serine 354 of FDH because replacement of this serine with alanine prevented this covalent modification. |
| GO:0051604 protein maturation | IDA PMID:21238436 Enzymatic properties of ALDH1L2, a mitochondrial 10-formylte... | MODIFY | Summary: AASDHPPT matures apo-ALDH1L2 by a defined serine phosphopantetheinylation, making the generic process term unnecessarily broad. Reason: Site-directed evidence identifies Ser375 as the modification site and shows that the modification enables enzyme activity. GO:0018070 peptidyl-serine phosphopantetheinylation is the appropriate specific replacement for GO:0051604. Proposed replacements: peptidyl-serine phosphopantetheinylation Supporting Evidence: PMID:21238436 Our results, however, demonstrate that PPT modifies mtFDH exclusively at Ser375. |
| GO:0005829 cytosol | IDA PMID:12815048 Cloning, expression, and characterization of a human 4'-phos... | ACCEPT | Summary: The founding human AASDHPPT study reports predominant expression in the cytosolic compartment. Reason: Although only the abstract is present in the local cache, it explicitly states cytosolic localization, and the experimental UniProt annotation is consistent with later structural work. There is no basis to second-guess the curator; the term also does not claim exclusive localization. Supporting Evidence: PMID:12815048 The human enzyme, which is expressed mainly in the cytosolic compartment in a wide range of tissues, is a 329-residue, monomeric protein. |
| GO:0008897 holo-[acyl-carrier-protein] synthase activity | IDA PMID:12815048 Cloning, expression, and characterization of a human 4'-phos... | ACCEPT | Summary: The founding biochemical study directly characterized human AASDHPPT as a broad-specificity phosphopantetheinyl transferase for human carrier proteins. Reason: The cached abstract explicitly reports transfer of the CoA-derived phosphopantetheine group to conserved serine residues in both cytosolic FASN ACP and mitochondrial ACP. That is the reaction defined by holo-[acyl-carrier-protein] synthase activity. Supporting Evidence: PMID:12815048 The enzyme is capable of transferring the 4'-phosphopantetheine moiety of coenzyme A to a conserved serine residue in both the acyl carrier protein domain of the human cytosolic multifunctional fatty acid synthase and the acyl carrier protein associated independently with human mitochondria. |
| GO:0070062 extracellular exosome | HDA PMID:19056867 Large-scale proteomics and phosphoproteomics of urinary exos... | KEEP AS NON CORE | Summary: AASDHPPT was detected in a large urinary-exosome proteome, supporting presence in that preparation but not an exosomal core function. Reason: The HDA annotation derives from LC-MS/MS of 1,132 urinary-exosome proteins. Such detection is valid localization evidence for the sampled material, but it does not show selective exosome loading, secretion, or activity there. Cytosolic phosphopantetheinyl transfer remains the established core biology. Supporting Evidence: PMID:19056867 Here, we used LC-MS/MS to profile the proteome of human urinary exosomes. |
| GO:0000287 magnesium ion binding | IDA PMID:18022563 Mechanism and substrate recognition of human holo ACP syntha... | ACCEPT | Summary: Structural and kinetic data directly establish catalytic Mg2+ binding by AASDHPPT. Reason: Mg2+ is resolved in the CoA-bound active site, its coordinating residues are identified, and mutations reduce Mg2+ affinity and catalytic turnover. This supports a specific cofactor-binding function rather than incidental metal association. Supporting Evidence: PMID:18022563 Magnesium is coordinated by the Ξ± and Ξ² groups of the CoA pyrophosphate, the carboxylates of Asp129 and Glu181, and one water molecule |
| GO:0005515 protein binding | IPI PMID:18022563 Mechanism and substrate recognition of human holo ACP syntha... | MARK AS OVER ANNOTATED | Summary: The structure directly demonstrates AASDHPPT binding the FASN ACP domain, but protein binding is a non-informative abstraction of substrate recognition during catalysis. Reason: This is a well-supported, physiologically coherent enzyme-substrate interface rather than a dubious interaction. Nevertheless, the binding is already intrinsic to holo-[acyl-carrier-protein] synthase activity, and no separate generic binding function should be inferred from it. Supporting Evidence: PMID:18022563 The interaction between the two proteins takes place on a large interface and is mostly of hydrophobic character. |
| GO:0008897 holo-[acyl-carrier-protein] synthase activity | IDA PMID:18022563 Mechanism and substrate recognition of human holo ACP syntha... | ACCEPT | Summary: CoA-, Mg2+-, and ACP-bound structures plus mutagenesis directly define the human holo-ACP synthase reaction. Reason: The study resolves AASDHPPT with its donor and acceptor substrates and validates catalytic residues kinetically. It is strong direct evidence for the core phosphopantetheinyl-transferase molecular function. Supporting Evidence: PMID:18022563 Besides the structural data observed in the complexes, the role of Glu181 and Lys185 as key acid/base catalysts is strongly supported by the observed significant loss of activity for the Glu181Gln, Glu181Ala, and Lys185Ala mutant proteins. |
| GO:0005829 cytosol | TAS Reactome:R-HSA-199202 | ACCEPT | Summary: Reactome places the FASN ACP phosphopantetheinylation reaction and its catalyst AASDHPPT in the cytosol. Reason: The pathway statement is consistent with direct biochemical fractionation and with the cytosolic FASN substrate. This annotation records the compartment of a curated human reaction and does not exclude other compartmental pools. Supporting Evidence: Reactome:R-HSA-199202 Cytosolic AASDHPPT (alpha-aminoadipic semialdehyde dehydrogenase-phosphopantetheinyl transferase) catalyzes the transfer of a phosphopantetheine moiety from coenzyme A to serine 2156 within the ACP domain of FAS (fatty acyl synthase). |
| GO:0005759 mitochondrial matrix | ISO PMID:41061852 Mitochondrial phosphopantetheinylation is required for oxida... | NEW | Summary: Proposed human mitochondrial-matrix localization transferred by orthology from mouse Aasdhppt. Reason: Endogenous mouse Aasdhppt was detected in mitochondrial fractions and showed matrix-like protease protection in C2C12 myoblasts; complementary split-Venus assays also placed the tested PPT construct in the matrix. This supports an ISO annotation for human AASDHPPT through the mouse ortholog Q9CQF6, not an IDA annotation on the human protein; endogenous human matrix localization remains to be tested. Propagation Review Root cause: NO FAILURE CORE Sources checked: UniProtKB:Q9CQF6 SUPPORTS TRANSFER The source is the experimentally tested mouse Aasdhppt ortholog in C2C12 myoblasts; the human annotation is an orthology transfer. Supporting Evidence: PMID:41061852 We detected endogenous PPT expression in both mitochondria and post-mitochondrial supernatant (PMS) (Fig. 3C). PMID:41061852 In agreement with our fractionation data in Fig. 3C, we observed a faint mitochondrial PPT band, suggesting matrix localization of mitochondrial PPT (Supplementary Fig. 3F). |
| GO:0031108 holo-[acyl-carrier-protein] biosynthetic process | ISO PMID:41061852 Mitochondrial phosphopantetheinylation is required for oxida... | NEW | Summary: Proposed human holo-NDUFAB1 biosynthetic-process annotation transferred by orthology from mouse Aasdhppt. Reason: Aasdhppt loss in mouse C2C12 myoblasts reduced holo- and octanoyl-NDUFAB1 and downstream mitochondrial fatty acid synthesis. This supports transfer of the immediate holo-ACP biosynthetic process to human AASDHPPT by ISO through mouse Q9CQF6; it is not direct human IDA evidence. Propagation Review Root cause: NO FAILURE CORE Sources checked: UniProtKB:Q9CQF6 SUPPORTS TRANSFER Mouse C2C12 loss-of-function and rescue experiments support transfer of the NDUFAB1/mtFAS process to the human ortholog. Supporting Evidence: PMID:41061852 From these data, we conclude that PPT is required for 4βPP modification of NDUFAB1 and downstream mtFAS activity [15]. |
Loading supporting contentβ¦
Download this section (compressed HTML)Q: Is endogenous human AASDHPPT imported into the mitochondrial matrix, and how does its cytosol-to-matrix partitioning vary across tissues and metabolic states?
Q: Is Q9NRN7-2 translated into a stable protein, and does it retain any catalytic activity or acquire a localization distinct from canonical Q9NRN7-1?
Q: Which proposed human acceptors, especially AASDH and DHRS2, are directly and physiologically phosphopantetheinylated by AASDHPPT?
Experiment: Endogenously tag AASDHPPT in human cells and combine matrix-resolved fractionation, protease protection, and proximity labeling with precise N-terminal targeting- sequence mutations. Quantify apo, holo, and acylated NDUFAB1, mtFAS products, lipoylation, respiratory-complex assembly, and oxygen consumption, with wild-type and matrix-targeted rescue controls.
Hypothesis: A small endogenous human AASDHPPT pool enters the mitochondrial matrix through its N-terminal targeting region and is required there for NDUFAB1 maturation and mtFAS.
Type: compartment-resolved human-cell genetics and mitochondrial biochemistry
Experiment: Measure isoform-specific RNA and protein at endogenous levels, then compare Q9NRN7-1 and Q9NRN7-2 stability, localization, CoA/Mg2+ binding, and activity on FASN ACP and NDUFAB1 in matched cell-rescue and purified-protein assays. Do not infer an isoform-specific compartment from transcript detection alone.
Hypothesis: Truncated AASDHPPT isoform 2 is unstable or catalytically inactive because it lacks the C-terminal portion containing key active-site residues.
Type: isoform-resolved expression and enzyme analysis
Experiment: Use an acute endogenous AASDHPPT degron followed by quantitative chemical proteomics and targeted mass spectrometry for candidate acceptor serines. Rescue with wild-type or catalytic-dead AASDHPPT and reconstitute positive targets with purified proteins to distinguish direct substrates from downstream changes.
Hypothesis: AASDH and DHRS2 are endogenous human AASDHPPT substrates with measurable, enzyme-dependent phosphopantetheine site occupancy.
Type: acute perturbation and site-resolved chemical proteomics
What is not known β curated, literature-grounded statements of the open unknowns (the inverse of core functions).
Gap: Endogenous human AASDHPPT has not been directly localized to the mitochondrial matrix, and the size, regulation, and physiological importance of a human matrix pool remain unknown.
OPEN BIOLOGYCURATION RESIDUAL_SUBGAP
What is known: Human fractionation and reviewed UniProt evidence establish a predominantly cytosolic pool, and purified human enzyme modifies mitochondrial ACP in vitro. Mouse C2C12 loss-of-function, localization, targeting-sequence, and rescue assays directly establish a matrix pool required for NDUFAB1 modification and mtFAS. The same mouse rescue system found reduced mtFAS rescue for five substitutions corresponding to human ClinVar variants, strengthening the human-allele relevance without directly demonstrating an endogenous human matrix pool. The corresponding human location and process assignments therefore remain supported by orthology rather than IDA.
Significance: Direct human evidence is needed to define the compartmental route by which AASDHPPT matures NDUFAB1 and to calibrate the strength of human matrix and mtFAS annotations.
What would resolve it: Endogenously tag AASDHPPT in human cells and combine quantitative fractionation, protease protection, matrix-directed proximity labeling, and selective disruption of its N-terminal targeting region. Measure NDUFAB1 4'-phosphopantetheinylation, mtFAS products, protein lipoylation, and respiration with matched rescue controls.
Provenance (the field's own admissions):
Gap: It is unknown whether human AASDHPPT isoform 2 produces a stable protein with any phosphopantetheinyl-transferase activity or whether it has a distinct cellular location.
OPEN BIOLOGY RESIDUAL_SUBGAP
What is known: UniProt catalogs two splice isoforms; isoform 2 changes residue 138 and lacks residues 139-309, including Glu181 and Lys185, which are required for efficient catalysis in the canonical protein. No compartment or core activity is therefore assigned by isoform.
Significance: Establishing whether isoform 2 is translated and functional would prevent a truncated, untested sequence from being conflated with the canonical enzyme in compartment-specific or catalytic interpretations.
What would resolve it: Quantify isoform-specific transcript and protein abundance, then compare endogenous-level Q9NRN7-1 and Q9NRN7-2 for stability, cytosolic and mitochondrial localization, CoA/Mg2+ binding, and phosphopantetheinylation of FASN ACP and NDUFAB1. Use purified proteins to distinguish intrinsic activity from import or stability effects.
Provenance (the field's own admissions):
Gap: The endogenous AASDHPPT dependence and site occupancy of proposed substrates such as AASDH and DHRS2 have not been established directly in human cells.
OPEN BIOLOGYCURATION RESIDUAL_SUBGAP
What is known: Human AASDHPPT activity is established directly for FASN ACP, NDUFAB1, ALDH1L1, and ALDH1L2. AASDH is supported by heterologous yeast complementation, whereas human-cell chemical proteomics describes the DHRS2 modification as potential.
Significance: Resolving this substrate boundary would separate physiological human targets from conserved-pathway or discovery-screen candidates and sharpen the scope of the enzyme's broad specificity.
What would resolve it: Acutely degrade endogenous AASDHPPT in human cells and quantify site-resolved phosphopantetheinylation of AASDH and DHRS2 alongside established substrates. Rescue with wild-type and catalytic-dead AASDHPPT and confirm candidate sites by targeted mass spectrometry and purified-protein reconstitution.
Provenance (the field's own admissions):
Loading supporting contentβ¦
Download this section (compressed HTML)Loading supporting contentβ¦
Download this section (compressed HTML)