ACAD9

Existing Annotations Review

GO Term	Evidence	Action	Reason
GO:0003995 acyl-CoA dehydrogenase activity	IBA GO_REF:0000033	ACCEPT	Summary: ACAD9 is a bona fide acyl-CoA dehydrogenase, belonging to the ACAD family and retaining catalytic activity for the alpha,beta-dehydrogenation of fatty acyl-CoA substrates. The IBA annotation is phylogenetically inferred and well-supported by direct biochemical data from multiple publications (PMID:16020546, PMID:34646991, PMID:38086790). This is an appropriate level of specificity for the IBA evidence code. Reason: ACAD9 has experimentally demonstrated acyl-CoA dehydrogenase activity. Purified ACAD9 shows dehydrogenation activity of 83 min-1 with palmitoyl-CoA as substrate (PMID:34646991). The IBA annotation at the general acyl-CoA dehydrogenase level is appropriate given phylogenetic support across the ACAD family. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN000744030 SUPPORTS TRANSFER The PAINT ancestral assertion is compatible with retained ACAD9 dehydrogenase chemistry, directly demonstrated in human protein. The target among descendant evidence is legitimate experimental grounding, not circularity. The ancestral tree and alignment were not independently reconstructed; no target-specific catalytic loss contradicts this broad IBA. Supporting Evidence: PMID:34646991 As purified, the wild-type ACAD9 protein has a dehydrogenation activity of 83 min-1, attributable to the FAD-containing holo-form of the protein PMID:16020546 Purified mature ACAD-9 had maximal activity with long-chain unsaturated acyl-CoAs as substrates (C16:1-, C18:1-, C18:2-, C22:6-CoA)
GO:0003995 acyl-CoA dehydrogenase activity	IEA GO_REF:0000002	ACCEPT	Summary: IEA annotation based on InterPro domain mapping. ACAD9 contains the acyl-CoA dehydrogenase active site signature (IPR006089) which correctly predicts this enzymatic activity. Consistent with direct experimental evidence. Reason: The InterPro-based IEA annotation is correct and consistent with experimental data showing ACAD9 possesses acyl-CoA dehydrogenase activity (PMID:16020546, PMID:34646991). This is a broader annotation that is subsumed by the IBA and IDA annotations but not incorrect. Propagation Review Root cause: NO FAILURE CORE Sources checked: InterPro:IPR006089 SUPPORTS TRANSFER The matched acyl-CoA dehydrogenase active-site signature supports the broad chemistry; human enzyme assays corroborate activity, without making every ACAD-family substrate transferable. Supporting Evidence: PMID:16020546 Purified mature ACAD-9 had maximal activity with long-chain unsaturated acyl-CoAs as substrates
GO:0004466 long-chain fatty acyl-CoA dehydrogenase activity	IEA GO_REF:0000120	ACCEPT	Summary: IEA annotation from ARBA/Rhea mapping based on the EC 1.3.8.8 catalytic activity. ACAD9 does have demonstrated long-chain acyl-CoA dehydrogenase activity (PMID:16020546), making this a valid annotation. Reason: ACAD9 has experimentally demonstrated long-chain fatty acyl-CoA dehydrogenase activity with substrate preference for C16-C22 chain lengths (PMID:16020546). The IEA annotation correctly captures this function and is consistent with the IDA annotation from BHF-UCL. Propagation Review Root cause: NO FAILURE CORE Sources checked: ARBA:ARBA00088474 UNRESOLVED The rule identifier is traced, but its detailed predicates were not inspected. Acceptance rests independently on the curated reaction and human substrate assays. RHEA:17721 SUPPORTS TRANSFER The cached UniProt catalytic record explicitly lists this long-chain acyl-CoA/ETF dehydrogenation reaction, consistent with C16 substrate assays. EC:1.3.8.8 SUPPORTS TRANSFER This long-chain acyl-CoA dehydrogenase EC assignment is recorded with experimental evidence in the cached human UniProt record. Supporting Evidence: PMID:16020546 Purified mature ACAD-9 had maximal activity with long-chain unsaturated acyl-CoAs as substrates (C16:1-, C18:1-, C18:2-, C22:6-CoA)
GO:0005743 mitochondrial inner membrane	IEA GO_REF:0000044	ACCEPT	Summary: IEA annotation based on UniProtKB/Swiss-Prot subcellular location mapping. ACAD9 is experimentally localized to the mitochondrial inner membrane, matrix side, as a peripheral membrane protein (PMID:16020546, PMID:20816094). Reason: Multiple experimental studies confirm ACAD9 localizes to the mitochondrial inner membrane. Submitochondrial fractionation showed ACAD9 is membrane-associated (PMID:16020546). The UniProt entry explicitly states "Mitochondrion inner membrane; Peripheral membrane protein; Matrix side" with evidence from PMID:16020546 and PMID:20816094. Propagation Review Root cause: NO FAILURE CORE Sources checked: UniProtKB-SubCell:SL-0168 SUPPORTS TRANSFER The inner-mitochondrial-membrane location maps to the cached UniProt statement specifying a peripheral protein on the matrix side; fractionation independently supports mitochondrial membrane association. Supporting Evidence: PMID:16020546 Submitochondrial fractionation studies found native ACAD-9 to be associated with the mitochondrial membrane
GO:0006631 fatty acid metabolic process	IEA GO_REF:0000117	MODIFY	Summary: The broad ARBA fatty-acid metabolic process is supported by ACAD9 catalysis, but fatty acid beta-oxidation more precisely describes the directly demonstrated process. Reason: Refine the existing process assertion to GO:0006635. ACAD9 itself catalyzes the acyl-CoA dehydrogenation step, and human HEK293 knockout/rescue experiments support its context-dependent contribution to fatty acid oxidation (PMID:16020546; PMID:25721401). This supplies the precise process used in the enzymatic core without adding a redundant NEW annotation. The original ARBA rule predicates remain uninspected; the refinement relies on primary human evidence. Propagation Review Root cause: TERM SCOPING PROBLEM Sources checked: ARBA:ARBA00028669 UNRESOLVED Detailed ARBA predicates were not inspected. The broad process is independently supported; human catalysis and knockout/rescue evidence justify refining its scope to fatty acid beta-oxidation. Proposed replacements: fatty acid beta-oxidation Supporting Evidence: PMID:16020546 Purified mature ACAD-9 had maximal activity with long-chain unsaturated acyl-CoAs as substrates (C16:1-, C18:1-, C18:2-, C22:6-CoA) PMID:25721401 both cell lines exhibited a 35–40% decrease in whole-cell palmitate oxidation
GO:0016491 oxidoreductase activity	IEA GO_REF:0000043	ACCEPT	Summary: IEA annotation based on UniProtKB keyword mapping (KW-0560, Oxidoreductase). ACAD9 is indeed an oxidoreductase. This is a very general term but is not incorrect for an IEA. Reason: ACAD9 is an oxidoreductase (acyl-CoA dehydrogenase, EC 1.3.8.7/1.3.8.8). While this is a very broad parent term, it is correctly applied via keyword mapping and more specific child terms are also annotated. Propagation Review Root cause: NO FAILURE CORE Sources checked: UniProtKB-KW:KW-0560 SUPPORTS TRANSFER The oxidoreductase keyword is compatible with experimentally measured FAD-dependent acyl-CoA dehydrogenation; it does not specify substrate chain length. Supporting Evidence: PMID:16020546 Purified mature ACAD-9 had maximal activity with long-chain unsaturated acyl-CoAs as substrates
GO:0016627 oxidoreductase activity, acting on the CH-CH group of donors	IEA GO_REF:0000002	ACCEPT	Summary: IEA annotation based on InterPro domain mapping. ACAD9 catalyzes alpha,beta-dehydrogenation of fatty acyl-CoA, which is an oxidoreductase reaction acting on CH-CH groups. This is a correct intermediate-level term between general oxidoreductase and specific acyl-CoA dehydrogenase activity. Reason: The acyl-CoA dehydrogenase reaction catalyzed by ACAD9 involves oxidation of the CH-CH bond at the alpha-beta position of fatty acyl-CoA substrates. This InterPro-derived term is accurate and at an appropriate level of specificity for automated annotation. Propagation Review Root cause: NO FAILURE CORE Sources checked: InterPro:IPR006091 SUPPORTS TRANSFER The matched middle domain supports ACAD-family chemistry, independently corroborated by human ACAD9 activity; it does not establish chain-length specificity. InterPro:IPR009075 SUPPORTS TRANSFER The matched C-terminal domain supports the ACAD fold. Experimental human dehydrogenase activity supplies the catalytic evidence beyond domain membership. InterPro:IPR009100 SUPPORTS TRANSFER The N-terminal/middle-domain superfamily match is compatible with the broad CH-CH oxidoreductase annotation, not a specific substrate class. InterPro:IPR013786 SUPPORTS TRANSFER The matched N-terminal ACAD domain is compatible with the broad chemistry; FAD-dependent activity is independently demonstrated in human ACAD9. InterPro:IPR036250 SUPPORTS TRANSFER The C-terminal ACAD-like superfamily match is compatible with the broad reaction class, without determining substrate chain length. InterPro:IPR037069 SUPPORTS TRANSFER The N-terminal ACAD superfamily match supports the structural family assignment; human biochemical assays establish retained enzyme activity. InterPro:IPR046373 SUPPORTS TRANSFER The middle-domain superfamily match is compatible with acyl-CoA oxidation, independently confirmed in human ACAD9. Supporting Evidence: PMID:16020546 Purified mature ACAD-9 had maximal activity with long-chain unsaturated acyl-CoAs as substrates
GO:0031966 mitochondrial membrane	IEA GO_REF:0000117	ACCEPT	Summary: IEA annotation from ARBA. ACAD9 is localized to the mitochondrial inner membrane. This term is broader (parent) than the more specific mitochondrial inner membrane annotations also present, but is not incorrect. Reason: ACAD9 is experimentally localized to the mitochondrial membrane (specifically the inner membrane). This broader IEA term is subsumed by more specific annotations but remains valid. Propagation Review Root cause: NO FAILURE CORE Sources checked: ARBA:ARBA00026364 UNRESOLVED Rule predicates were not inspected. Independent submitochondrial fractionation in PMID:16020546 supports mitochondrial membrane association. Supporting Evidence: PMID:16020546 Submitochondrial fractionation studies found native ACAD-9 to be associated with the mitochondrial membrane
GO:0050660 flavin adenine dinucleotide binding	IEA GO_REF:0000002	ACCEPT	Summary: IEA annotation based on InterPro domain mapping. ACAD9 is a FAD-containing flavoprotein. FAD binding has been experimentally demonstrated, with approximately 70% FAD occupancy in purified protein (PMID:34646991). FAD binding is essential for the dehydrogenase function, and ECSIT binding causes deflavination (PMID:33320993, PMID:38086790). Reason: ACAD9 binds FAD as a cofactor for its acyl-CoA dehydrogenase activity. The FAD content of purified ACAD9 is approximately 70% (PMID:34646991), and ECSIT-mediated deflavination switches ACAD9 from FAO enzyme to CI assembly factor (PMID:33320993). Propagation Review Root cause: NO FAILURE CORE Sources checked: InterPro:IPR013786 SUPPORTS TRANSFER The matched ACAD-family domain is compatible with FAD binding; ACAD9 cofactor content, deflavination, and reconstitution were directly measured in PMID:33320993 and PMID:34646991. InterPro:IPR037069 SUPPORTS TRANSFER The matched ACAD-family domain is compatible with FAD binding; ACAD9 cofactor content, deflavination, and reconstitution were directly measured in PMID:33320993 and PMID:34646991. Supporting Evidence: PMID:34646991 the FAD content of purified wild type ACAD9 protein (non-tagged ACAD9 and ACAD9-His6) was approximately 70% PMID:33320993 interaction with ECSIT induces ACAD9 to eject its FAD cofactor from the catalytic site
GO:0070991 medium-chain fatty acyl-CoA dehydrogenase activity	IEA GO_REF:0000120	KEEP AS NON CORE	Summary: The combined electronic mapping traces to RHEA:14477 and EC:1.3.8.7, both present in the curated ACAD9 catalytic record. Medium-chain activity is retained as secondary to the established long-chain reaction. Reason: The cached human UniProt record assigns medium-chain chemistry and individual C9-C11 reactions to PMID:16020546. Its primary substrate panel could not be independently recovered, so this judgment defers to that experimental curation and does not claim fresh verification of the C9-C11 measurements. The long-chain preference is explicit in the accessible abstract. No broad physiological medium-chain oxidation role is inferred. Propagation Review Root cause: NO FAILURE NON CORE Sources checked: RHEA:14477 SUPPORTS TRANSFER The cached UniProt record lists this medium-chain acyl-CoA dehydrogenation reaction with PMID:16020546 evidence; the exact primary substrate panel was not recovered in this audit. EC:1.3.8.7 SUPPORTS TRANSFER The experimentally curated EC assignment and individual nonanoyl/decanoyl/undecanoyl reactions support retaining a secondary activity; this does not establish a major physiological medium-chain FAO role. Supporting Evidence: PMID:16020546 Purified mature ACAD-9 had maximal activity with long-chain unsaturated acyl-CoAs as substrates (C16:1-, C18:1-, C18:2-, C22:6-CoA)
GO:0017099 very-long-chain fatty acyl-CoA dehydrogenase activity	IDA PMID:33320993 Assembly of The Mitochondrial Complex I Assembly Complex Sug...	MODIFY	Summary: PMID:33320993 demonstrates ACAD9 dehydrogenation using palmitoyl-CoA (C16:0) in a ETF fluorescence reduction assay. This supports long-chain fatty acyl-CoA dehydrogenase activity. Reason: The source assay measures a C16 substrate. The verified GO definitions distinguish long-chain substrates (13-22 carbons; GO:0004466) from very-long-chain substrates (more than 22 carbons; GO:0017099). Replace the term with the activity actually demonstrated by this source. This is an evidence-scope correction, not proof that ACAD9 can never use longer substrates. Relative activity versus the protein named VLCAD and an unvalidated substrate-channel prediction do not determine this GO chain-length classification. Comparator ACADVL also carries GO:0017099 in its cached curated record, including IDA PMID:9461620, whose abstract describes palmitoyl-CoA assays. This reveals a possible difference between enzyme-class usage and the explicit chain-length definition; the present replacement follows the demonstrated ACAD9 substrate and does not assert that the comparator annotation is erroneous. Proposed replacements: long-chain fatty acyl-CoA dehydrogenase activity Supporting Evidence: PMID:33320993 After addition of the ACAD specific substrate palmitoyl-CoA (C16:0), there is a clear loss of ETF fluorescence in ACAD9 alone PMID:16020546 Purified mature ACAD-9 had maximal activity with long-chain unsaturated acyl-CoAs as substrates (C16:1-, C18:1-, C18:2-, C22:6-CoA)
GO:0017099 very-long-chain fatty acyl-CoA dehydrogenase activity	IDA PMID:34646991 Molecular mechanism of interactions between ACAD9 and bindin...	MODIFY	Summary: PMID:34646991 demonstrates ACAD9 dehydrogenation using palmitoyl-CoA (C16:0) in a ferricenium reduction assay. This supports long-chain fatty acyl-CoA dehydrogenase activity. Reason: The source assay measures a C16 substrate. The verified GO definitions distinguish long-chain substrates (13-22 carbons; GO:0004466) from very-long-chain substrates (more than 22 carbons; GO:0017099). Replace the term with the activity actually demonstrated by this source. This is an evidence-scope correction, not proof that ACAD9 can never use longer substrates. Relative activity versus the protein named VLCAD and an unvalidated substrate-channel prediction do not determine this GO chain-length classification. Comparator ACADVL also carries GO:0017099 in its cached curated record, including IDA PMID:9461620, whose abstract describes palmitoyl-CoA assays. This reveals a possible difference between enzyme-class usage and the explicit chain-length definition; the present replacement follows the demonstrated ACAD9 substrate and does not assert that the comparator annotation is erroneous. Proposed replacements: long-chain fatty acyl-CoA dehydrogenase activity Supporting Evidence: PMID:34646991 Reactions contained 0.1 μM purified ACAD9 or VLCAD protein, 50 μM palmitoyl-CoA, and 200 μM ferricenium hexafluorophosphate
GO:0030674 protein-macromolecule adaptor activity	IDA PMID:32320651 Dissecting the Roles of Mitochondrial Complex I Intermediate...	UNDECIDED	Summary: The 2020 study establishes ACAD9-dependent MCIA stability and assembly, but the precise adaptor mechanism assigned by this IDA remains unresolved. Reason: The full 2020 paper was inspected through the author-institution PDF: it establishes ACAD9-dependent MCIA integrity and assembly-intermediate association. These results do not identify two distinct macromolecules directly bridged by ACAD9. In PMID:33320993, ECSIT is the experimentally resolved bridge and no direct ACAD9-NDUFAF1 interaction was observed. GO:0030674 requires bringing macromolecules together; MCIA membership alone does not settle that specific mechanism. Preserve the curator assertion as UNDECIDED without declaring it erroneous or substituting a scaffold function. Complex membership and participation in complex I assembly remain established. Supporting Evidence: PMID:32320651 while each MCIA component is critical for complex I assembly, a hierarchy of stability exists centered on ACAD9 PMID:33320993 No direct interaction between NDUFAF1 and ACAD9 was observed
GO:0160295 mitochondrial complex I intermediate assembly complex	IDA PMID:33320993 Assembly of The Mitochondrial Complex I Assembly Complex Sug...	ACCEPT	Summary: IDA annotation from FlyBase based on Giachin et al. 2021. This study provided extensive structural and biochemical evidence that ACAD9 is a core component of the MCIA complex, including cryo-EM structures, SAXS, native MS, and SEC-MALLS data demonstrating stable ACAD9-ECSIT complexes. Reason: ACAD9 is unequivocally a component of the MCIA complex. Giachin et al. demonstrated formation of the ACAD9-ECSIT binary complex by multiple biophysical methods and showed ECSIT binds the vestigial dehydrogenase domain of ACAD9 (PMID:33320993). This is a core annotation for ACAD9. Supporting Evidence: PMID:33320993 ECSIT functions as the bridging node of the MCIA core complex PMID:33320993 the C-terminal domain of ECSIT directly binds to the vestigial dehydrogenase domain of the FAO enzyme ACAD9 and induces its deflavination
GO:0160295 mitochondrial complex I intermediate assembly complex	IDA PMID:34646991 Molecular mechanism of interactions between ACAD9 and bindin...	ACCEPT	Summary: IDA annotation from FlyBase based on Xia et al. 2021 (iScience). This study reconstituted the ACAD9/ECSIT/NDUFAF1 ternary complex in vitro and characterized it by SEC, SAXS, and pull-down assays, demonstrating ACAD9 is a core member of the MCIA complex. Reason: Xia et al. provided direct biochemical evidence for ACAD9 as part of the MCIA ternary complex with ECSIT and NDUFAF1, including purification of the stable ternary complex with 1:1:1 stoichiometry (PMID:34646991). This is strong supporting evidence for the CC annotation. Supporting Evidence: PMID:34646991 ACAD9, ECSIT, and NDUFAF1 form the core mitochondrial CI assembly (MCIA) complex PMID:34646991 mixing of the three purified, individual proteins forms a ternary complex, ACAD9/ECSIT/NDUFAF1, with a 1:1:1 molar (monomer) ratio
GO:0005739 mitochondrion	IDA GO_REF:0000052	ACCEPT	Summary: IDA annotation from HPA based on immunofluorescence data. ACAD9 localization to mitochondria is well-established across multiple studies and methods. Reason: ACAD9 mitochondrial localization is confirmed by multiple independent approaches: submitochondrial fractionation (PMID:16020546), immunofluorescence (PMID:20816094), and proteomics (PMID:34800366). The HPA immunofluorescence data is consistent with this body of evidence. Supporting Evidence: PMID:16020546 Submitochondrial fractionation studies found native ACAD-9 to be associated with the mitochondrial membrane
GO:0160295 mitochondrial complex I intermediate assembly complex	IDA PMID:32320651 Dissecting the Roles of Mitochondrial Complex I Intermediate...	ACCEPT	Summary: IDA annotation from FlyBase based on Formosa et al. 2020. This study used cell knockout studies of each MCIA component and demonstrated that ACAD9 is central to MCIA complex stability, also identifying TMEM186 and COA1 as additional MCIA components. Reason: Formosa et al. provided strong genetic evidence from knockout studies that ACAD9 is a core component of the MCIA complex, with a central role in maintaining complex stability. Loss of ACAD9 resulted in destabilization of the entire MCIA complex and impaired CI assembly (PMID:32320651). Supporting Evidence: PMID:32320651 The mitochondrial complex I intermediate assembly (MCIA) complex, containing assembly factors NDUFAF1, ECSIT, ACAD9, and TMEM126B, is required for building the intermediate ND2-module PMID:32320651 while each MCIA component is critical for complex I assembly, a hierarchy of stability exists centered on ACAD9
GO:0003995 acyl-CoA dehydrogenase activity	IDA PMID:38086790 The assembly of the Mitochondrial Complex I Assembly complex...	ACCEPT	Summary: IDA annotation from FlyBase based on McGregor et al. 2023 (Nature Communications). This study determined a 3.0 A cryo-EM structure of the ACAD9-ECSIT complex and also characterized ACAD9 dehydrogenase activity using the ETF fluorescence reduction assay with palmitoyl-CoA. The study demonstrated ECSIT binding induces a large conformational change in the FAD-binding loop of ACAD9, releasing FAD and converting ACAD9 from an FAO enzyme to a CI assembly factor. Reason: McGregor et al. directly measured ACAD9 acyl-CoA dehydrogenase activity via ETF fluorescence reduction assay and confirmed the deflavination mechanism (PMID:38086790). This provides direct experimental evidence for ACAD9's enzymatic function. Supporting Evidence: PMID:38086790 ECSIT binding induces a major conformational change in the FAD-binding loop of ACAD9, releasing the FAD cofactor and converting ACAD9 from a fatty acid beta-oxidation (FAO) enzyme to a CI assembly factor
GO:0005743 mitochondrial inner membrane	NAS PMID:32320651 Dissecting the Roles of Mitochondrial Complex I Intermediate...	ACCEPT	Summary: NAS annotation from ComplexPortal based on PMID:32320651. ACAD9 functions as part of the MCIA complex at the mitochondrial inner membrane where CI assembly occurs. This is consistent with the experimentally determined localization. Reason: ACAD9 is localized to the mitochondrial inner membrane, matrix side, as a peripheral membrane protein. This is well-supported by fractionation studies (PMID:16020546) and is the site of its CI assembly function as part of the MCIA complex (PMID:32320651). Supporting Evidence: PMID:16020546 Submitochondrial fractionation studies found native ACAD-9 to be associated with the mitochondrial membrane
GO:0032981 mitochondrial respiratory chain complex I assembly	NAS PMID:32320651 Dissecting the Roles of Mitochondrial Complex I Intermediate...	ACCEPT	Summary: NAS annotation from ComplexPortal based on Formosa et al. 2020. ACAD9 is essential for CI assembly as a core member of the MCIA complex. This is the primary biological process annotation for ACAD9 and represents its most important physiological function. Reason: Complex I assembly is a direct ACAD9 function: the source identifies it as a core MCIA component required for assembly of the ND2-module. The cell-knockout results establish assembly dependence without relying on an unverified claim about where disease variants cluster. The distinct dehydrogenase function does not weaken this assembly annotation. Supporting Evidence: PMID:32320651 The mitochondrial complex I intermediate assembly (MCIA) complex, containing assembly factors NDUFAF1, ECSIT, ACAD9, and TMEM126B, is required for building the intermediate ND2-module PMID:20816094 ACAD9 binds complex I assembly factors NDUFAF1 and Ecsit and is specifically required for the assembly of complex I file:human/ACAD9/ACAD9-deep-research-falcon.md its dominant essential role is as a CI assembly factor within MCIA.
GO:0005739 mitochondrion	HTP PMID:34800366 Quantitative high-confidence human mitochondrial proteome an...	ACCEPT	Summary: The HTP source assigns ACAD9 to mitochondria in a proteome-scale study. This location is independently established by targeted ACAD9 experiments. Reason: Accept mitochondrial localization with curator deference and independent fractionation evidence in PMID:16020546. The publication describes the MitoCoP workflow, but its ACAD9-specific supplementary row was not separately re-extracted here; the generic proteome-size statement is not treated as direct identification of ACAD9. Supporting Evidence: PMID:16020546 Submitochondrial fractionation studies found native ACAD-9 to be associated with the mitochondrial membrane
GO:0005515 protein binding	IPI PMID:33753518 TMEM70 and TMEM242 help to assemble the rotor ring of human ...	REMOVE	Summary: Affinity enrichment of tagged TMEM70 and TMEM242 in human HEK293 cells recovered ACAD9 and other MCIA proteins. The generic protein-binding term does not identify ACAD9 molecular function. Reason: Remove the uninformative generic term under the binding curation policy, without denying the reported co-association. The enrichment experiment does not by itself establish direct pairwise binding or a specific adaptor activity of ACAD9. MCIA membership and assembly function are retained in their independently supported annotations. Supporting Evidence: PMID:33753518 TMEM70 and TMEM242 interact with the mitochondrial complex I assembly (the MCIA) complex that supports assembly of the membrane arm of complex I
GO:0005515 protein binding	IPI PMID:32320651 Dissecting the Roles of Mitochondrial Complex I Intermediate...	REMOVE	Summary: The 2020 assembly study supports ACAD9 association with MCIA components. Protein binding provides no specific molecular-function description. Reason: Remove the uninformative generic annotation without rejecting the underlying interactions. The complex-stability and association results support the established MCIA role but do not independently resolve a direct ACAD9 adaptor mechanism, so no new molecular-function replacement is inferred from this IPI row. Supporting Evidence: PMID:32320651 The mitochondrial complex I intermediate assembly (MCIA) complex, containing assembly factors NDUFAF1, ECSIT, ACAD9, and TMEM126B, is required for building the intermediate ND2-module
GO:0001676 long-chain fatty acid metabolic process	IDA PMID:16020546 Human acyl-CoA dehydrogenase-9 plays a novel role in the mit...	ACCEPT	Summary: ACAD9 directly catalyzes long-chain acyl-CoA dehydrogenation, including unsaturated substrates. This is a supported biochemical function alongside complex I assembly. Reason: The source establishes long-chain substrate catalysis. Human knockout/rescue results in PMID:25721401 additionally demonstrate a cellular long-chain FAO contribution. Retain this process as core to the enzymatic function; the existence of an essential assembly role does not make all long-chain metabolism incidental. Earlier disease observations without detectable FAO abnormalities should not be generalized to every tissue or variant. Supporting Evidence: PMID:16020546 Purified mature ACAD-9 had maximal activity with long-chain unsaturated acyl-CoAs as substrates (C16:1-, C18:1-, C18:2-, C22:6-CoA) PMID:25721401 both cell lines exhibited a 35–40% decrease in whole-cell palmitate oxidation and palmitoyl-CoA ACAD activity
GO:0004466 long-chain fatty acyl-CoA dehydrogenase activity	IDA PMID:16020546 Human acyl-CoA dehydrogenase-9 plays a novel role in the mit...	ACCEPT	Summary: Purified ACAD9 displays maximal activity on the tested unsaturated C16:1-, C18:1-, C18:2-, and C22:6-CoA substrates, supporting long-chain fatty acyl-CoA dehydrogenase activity. Reason: The accessible primary abstract explicitly reports these substrates, all within the current GO long-chain definition of 13-22 carbons. Independent C16 assays in PMID:33320993 and PMID:34646991 corroborate the activity. Precise kinetic constants are omitted because the original kinetic table was not independently recovered. Supporting Evidence: PMID:16020546 Purified mature ACAD-9 had maximal activity with long-chain unsaturated acyl-CoAs as substrates (C16:1-, C18:1-, C18:2-, C22:6-CoA)
GO:0005739 mitochondrion	IDA PMID:16020546 Human acyl-CoA dehydrogenase-9 plays a novel role in the mit...	ACCEPT	Summary: IDA annotation from BHF-UCL based on Ensenauer et al. 2005. Submitochondrial fractionation studies confirmed ACAD9 is associated with the mitochondrial membrane. Reason: Direct experimental evidence from submitochondrial fractionation demonstrates ACAD9 is a mitochondrial protein associated with mitochondrial membranes (PMID:16020546). Supporting Evidence: PMID:16020546 Submitochondrial fractionation studies found native ACAD-9 to be associated with the mitochondrial membrane
GO:0031966 mitochondrial membrane	IDA PMID:16020546 Human acyl-CoA dehydrogenase-9 plays a novel role in the mit...	ACCEPT	Summary: IDA annotation from BHF-UCL based on Ensenauer et al. 2005. Submitochondrial fractionation showed ACAD9 is membrane-associated. This is less specific than the inner membrane annotation but still correct based on the fractionation data. Reason: ACAD9 is a peripheral membrane protein associated with the mitochondrial inner membrane on the matrix side (PMID:16020546). The mitochondrial membrane annotation is correct, though the more specific inner membrane annotation is also present. Supporting Evidence: PMID:16020546 Submitochondrial fractionation studies found native ACAD-9 to be associated with the mitochondrial membrane
GO:0051791 medium-chain fatty acid metabolic process	IDA PMID:16020546 Human acyl-CoA dehydrogenase-9 plays a novel role in the mit...	KEEP AS NON CORE	Summary: The experimental annotation records medium-chain activity of ACAD9 in PMID:16020546. The curated UniProt reaction list includes C9-C11 substrates, while the accessible primary abstract emphasizes long-chain preference. Reason: Retain as non-core with deference to the experimental curator and the concordant curated catalytic record. The original medium-chain substrate panel was not independently recovered; no quantitative comparison with MCAD is asserted. These data do not establish medium-chain oxidation as a principal physiological ACAD9 function. Supporting Evidence: PMID:16020546 Purified mature ACAD-9 had maximal activity with long-chain unsaturated acyl-CoAs as substrates (C16:1-, C18:1-, C18:2-, C22:6-CoA)
GO:0070991 medium-chain fatty acyl-CoA dehydrogenase activity	IDA PMID:16020546 Human acyl-CoA dehydrogenase-9 plays a novel role in the mit...	KEEP AS NON CORE	Summary: The experimental annotation records medium-chain activity of ACAD9 in PMID:16020546. The curated UniProt reaction list includes C9-C11 substrates, while the accessible primary abstract emphasizes long-chain preference. Reason: Retain as non-core with deference to the experimental curator and the concordant curated catalytic record. The original medium-chain substrate panel was not independently recovered; no quantitative comparison with MCAD is asserted. These data do not establish medium-chain oxidation as a principal physiological ACAD9 function. Supporting Evidence: PMID:16020546 Purified mature ACAD-9 had maximal activity with long-chain unsaturated acyl-CoAs as substrates (C16:1-, C18:1-, C18:2-, C22:6-CoA)
GO:0005743 mitochondrial inner membrane	TAS Reactome:R-HSA-6799179	ACCEPT	Summary: TAS annotation from Reactome pathway "Peripheral arm subunits bind the 815kDa complex to form a 980kDa complex". ACAD9 functions at the mitochondrial inner membrane as part of the MCIA complex during CI biogenesis. Reason: ACAD9 is localized to the mitochondrial inner membrane where CI assembly takes place. This is well-supported experimentally (PMID:16020546, PMID:20816094) and the Reactome annotation correctly reflects this. Supporting Evidence: PMID:16020546 Submitochondrial fractionation studies found native ACAD-9 to be associated with the mitochondrial membrane
GO:0005743 mitochondrial inner membrane	TAS Reactome:R-HSA-6799196	ACCEPT	Summary: TAS annotation from Reactome pathway "The MCIA complex, NDUFAF2-7 all dissociate from the 980kDa complex, resulting in Complex I". Consistent with ACAD9's role as an MCIA assembly factor at the inner membrane. Reason: Same as above. ACAD9 is at the mitochondrial inner membrane and this Reactome step describes MCIA complex dissociation after CI assembly is complete. Well-supported experimentally. Supporting Evidence: PMID:16020546 Submitochondrial fractionation studies found native ACAD-9 to be associated with the mitochondrial membrane
GO:0005743 mitochondrial inner membrane	TAS Reactome:R-HSA-6799197	ACCEPT	Summary: TAS annotation from Reactome pathway "ND4, ND5 bind the 550kDa complex to form the 815kDa complex". Another step in CI biogenesis at the inner membrane. Reason: Consistent with ACAD9's role as an MCIA assembly factor at the mitochondrial inner membrane. Supporting Evidence: PMID:16020546 Submitochondrial fractionation studies found native ACAD-9 to be associated with the mitochondrial membrane
GO:0005743 mitochondrial inner membrane	TAS Reactome:R-HSA-6799199	ACCEPT	Summary: TAS annotation from Reactome pathway "COA1:MT-ND2, TMEM186:MT-ND3, MT-ND6, NDUFB6 bind the MCIA complex to form a 370kDa subcomplex". This step directly involves the MCIA complex of which ACAD9 is a core member. Reason: ACAD9 is a core member of the MCIA complex that participates in this assembly step. Well-supported by PMID:32320651. Supporting Evidence: PMID:32320651 The mitochondrial complex I intermediate assembly (MCIA) complex, containing assembly factors NDUFAF1, ECSIT, ACAD9, and TMEM126B, is required for building the intermediate ND2-module
GO:0005743 mitochondrial inner membrane	TAS Reactome:R-HSA-6799202	ACCEPT	Summary: TAS annotation from Reactome pathway "The 315kDa subcomplex binds the 370kDa subcomplex to form the 550kDa complex". Another CI assembly step at the inner membrane. Reason: Consistent with ACAD9's role as an MCIA assembly factor at the mitochondrial inner membrane. Supporting Evidence: PMID:16020546 Submitochondrial fractionation studies found native ACAD-9 to be associated with the mitochondrial membrane
GO:0005515 protein binding	IPI PMID:20816094 Acyl-CoA dehydrogenase 9 is required for the biogenesis of o...	REMOVE	Summary: The source explicitly reports ACAD9 association with NDUFAF1 and ECSIT and a requirement for complex I assembly. Generic protein binding is uninformative. Reason: Remove the generic term under the binding policy while retaining the reported association and the separately annotated assembly function. The accessible source abstract supports the interaction assertion, so this is not a judgment that the interaction is false. Later reconstitution resolves ECSIT as the direct bridging component; association alone does not warrant a new ACAD9 adaptor assignment. Supporting Evidence: PMID:20816094 ACAD9 binds complex I assembly factors NDUFAF1 and Ecsit and is specifically required for the assembly of complex I
GO:0005739 mitochondrion	IDA PMID:20816094 Acyl-CoA dehydrogenase 9 is required for the biogenesis of o...	ACCEPT	Summary: IDA annotation based on Nouws et al. 2010. ACAD9 was shown to localize to mitochondria as part of the study establishing its role in CI assembly. Reason: ACAD9 mitochondrial localization is well-established and confirmed in this study (PMID:20816094) which demonstrated ACAD9 interactions with mitochondrial CI assembly factors. Supporting Evidence: PMID:20816094 ACAD9 binds complex I assembly factors NDUFAF1 and Ecsit and is specifically required for the assembly of complex I
GO:0032981 mitochondrial respiratory chain complex I assembly	IMP PMID:20816094 Acyl-CoA dehydrogenase 9 is required for the biogenesis of o...	ACCEPT	Summary: The source demonstrates ACAD9 participation in complex I biogenesis through interaction and perturbation studies. Reason: Complex I assembly is an established direct role of ACAD9, supported by the source and later MCIA studies. The absence of disturbed long-chain FAO reported in the 2010 study applies to its tested models; PMID:25721401 demonstrates long-chain FAO involvement in ACAD9-deficient HEK293 cells. That contextual difference does not weaken the complex I assembly annotation. Supporting Evidence: PMID:20816094 ACAD9 binds complex I assembly factors NDUFAF1 and Ecsit and is specifically required for the assembly of complex I PMID:20816094 ACAD9 mutations result in complex I deficiency and not in disturbed long-chain fatty acid oxidation
GO:0005634 nucleus	IDA PMID:21237683 Identification and characterization of new long chain acyl-C...	UNDECIDED	Summary: The cellular-nucleus annotation remains unresolved after reading the abstract-only local cache and externally accessed sections of PMC3073726. Reason: Externally accessed PMC3073726 section 3.8 (https://pmc.ncbi.nlm.nih.gov/articles/PMC3073726/) discusses ACAD11 nuclear staining and ACAD9 staining in the anatomical dentate nucleus, which is distinct from GO:0005634. The local PMID:21237683 abstract only describes granular-layer expression; that statement does not substantiate a cellular nuclear pool. Complete ACAD9-specific evidence supporting the nuclear curator assignment remains unresolved, so retain UNDECIDED without asserting misattribution or using mitochondrial targeting to exclude a nuclear pool.
GO:0030425 dendrite	IDA PMID:21237683 Identification and characterization of new long chain acyl-C...	KEEP AS NON CORE	Summary: Externally accessed PMC3073726 section 3.8 reports ACAD9 immunostaining in dendrites of granular neurons in human cerebellum; the local PMID:21237683 cache contains only the abstract. Reason: Retain this observed neuronal compartment as non-core based on the externally read primary section (https://pmc.ncbi.nlm.nih.gov/articles/PMC3073726/, section 3.8) and the existing IDA curator assignment. The cached granular-layer statement alone does not establish dendrites and is not used as the supporting quote for this decision. Dendritic localization is compatible with mitochondrial association but does not establish a separate dendrite-specific molecular activity.
GO:0005739 mitochondrion	IDA GO_REF:0000054	ACCEPT	Summary: IDA annotation from LIFEdb based on expressed fusion protein localization in living cells. ACAD9 localization to mitochondria was confirmed, consistent with all other evidence. Reason: ACAD9 mitochondrial localization is well-established. The LIFEdb fusion protein experiment confirms what is known from multiple other approaches (PMID:16020546, PMID:20816094, PMID:34800366). Supporting Evidence: PMID:16020546 Submitochondrial fractionation studies found native ACAD-9 to be associated with the mitochondrial membrane

Gene Description

Existing Annotations Review

Core Functions

Promotes mitochondrial complex I assembly as a core MCIA component required for the stability of other assembly factors and formation of the ND2-module at the inner membrane. ACAD9 binds ECSIT; ECSIT connects it to NDUFAF1 and induces deflavination.

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