id: P40939
gene_symbol: HADHA
product_type: PROTEIN
status: COMPLETE
taxon:
  id: NCBITaxon:9606
  label: Homo sapiens
description: >-
  HADHA encodes the alpha subunit of the mitochondrial trifunctional protein (MTP/TFP),
  a heterotetrameric complex (alpha2-beta2) with HADHB that is bound to the matrix face of
  the mitochondrial inner membrane and catalyzes the last three steps of long-chain fatty
  acid beta-oxidation. The alpha subunit carries TWO of these activities: long-chain
  enoyl-CoA hydratase (LCEH, EC 4.2.1.17), which hydrates 2-trans-enoyl-CoA to
  (3S)-3-hydroxyacyl-CoA, and long-chain 3-hydroxyacyl-CoA dehydrogenase (LCHAD, EC
  1.1.1.211), which oxidizes (3S)-3-hydroxyacyl-CoA to 3-oxoacyl-CoA using NAD+. The third
  step (thiolytic cleavage to acetyl-CoA) is carried by the beta subunit (HADHB). The
  enzyme is specific for long-chain (C10-C16) acyl-CoA substrates, including saturated and
  unsaturated species. Independently of the beta subunit, the alpha subunit also exhibits a
  monolysocardiolipin acyltransferase activity that participates in cardiolipin remodeling.
  Deficiency of HADHA causes isolated LCHAD deficiency and generalized mitochondrial
  trifunctional protein deficiency, presenting with hypoketotic hypoglycemia,
  cardiomyopathy, peripheral neuropathy and pigmentary retinopathy, and associated with
  maternal HELLP syndrome / acute fatty liver of pregnancy.
alternative_products:
- name: '1'
  id: P40939-1
- name: '2'
  id: P40939-2
  sequence_note: VSP_059010, VSP_059011
existing_annotations:
# ============================================================================
# MOLECULAR FUNCTION - ENOYL-CoA HYDRATASE (CORE - alpha subunit)
# ============================================================================
- term:
    id: GO:0004300
    label: enoyl-CoA hydratase activity
  evidence_type: IBA
  original_reference_id: GO_REF:0000033
  qualifier: enables
  review:
    summary: >-
      Phylogenetic (IBA) annotation assigning enoyl-CoA hydratase activity (EC 4.2.1.17) to
      HADHA. This is one of the two correct core catalytic activities of the alpha subunit:
      it hydrates 2-trans-enoyl-CoA to (3S)-3-hydroxyacyl-CoA in step 2 of each
      beta-oxidation cycle. Directly demonstrated for the human alpha subunit by expression
      studies (PMID:8135828) and confirmed structurally (PMID:30850536).
    action: ACCEPT
    reason: >-
      Core molecular function of HADHA, well supported by direct expression and structural
      evidence.
    supported_by:
    - reference_id: PMID:8135828
      supporting_text: "Expression of this cDNA in mammalian cells yielded a polypeptide with
        the long-chain enoyl-CoA hydratase and long-chain 3-hydroxyacyl-CoA dehydrogenase
        activities."
      full_text_unavailable: true
- term:
    id: GO:0004300
    label: enoyl-CoA hydratase activity
  evidence_type: IEA
  original_reference_id: GO_REF:0000120
  qualifier: enables
  review:
    summary: >-
      Electronic (IEA) annotation of enoyl-CoA hydratase activity based on InterPro/EC
      mapping. Consistent with the IBA and direct experimental evidence.
    action: ACCEPT
    reason: >-
      Redundant with the experimental annotation but correctly captures the core hydratase
      function of the alpha subunit.
- term:
    id: GO:0004300
    label: enoyl-CoA hydratase activity
  evidence_type: EXP
  original_reference_id: PMID:1550553
  qualifier: enables
  review:
    summary: >-
      Experimental annotation of enoyl-CoA hydratase activity. PMID:1550553 purified the
      human liver multifunctional enzyme and showed it possesses 2-enoyl-CoA hydratase
      activity in addition to dehydrogenase and thiolase. Although this early paper did not
      separate subunit-specific activities, PMID:8135828 subsequently localized the hydratase
      to the alpha subunit (HADHA).
    action: ACCEPT
    reason: >-
      Core molecular function with direct experimental support; subunit assignment confirmed
      by PMID:8135828.
    supported_by:
    - reference_id: PMID:1550553
      supporting_text: "the enzyme possesses 2-enoyl-CoA hydratase and 3-ketoacyl-CoA thiolase
        activities which cannot be separated from the dehydrogenase."
      full_text_unavailable: true
- term:
    id: GO:0004300
    label: enoyl-CoA hydratase activity
  evidence_type: TAS
  original_reference_id: PMID:8135828
  qualifier: enables
  review:
    summary: >-
      TAS annotation of enoyl-CoA hydratase activity citing PMID:8135828, the paper that
      definitively showed expression of the alpha-subunit cDNA yields hydratase (and
      dehydrogenase) activity.
    action: ACCEPT
    reason: >-
      Core molecular function with strong direct evidence assigning the hydratase to the
      alpha subunit specifically.
    supported_by:
    - reference_id: PMID:8135828
      supporting_text: "Expression of this cDNA in mammalian cells yielded a polypeptide with
        the long-chain enoyl-CoA hydratase and long-chain 3-hydroxyacyl-CoA dehydrogenase
        activities."
      full_text_unavailable: true
# ============================================================================
# MOLECULAR FUNCTION - 3-HYDROXYACYL-CoA DEHYDROGENASE (CORE - alpha subunit)
# ============================================================================
- term:
    id: GO:0016509
    label: long-chain (3S)-3-hydroxyacyl-CoA dehydrogenase (NAD+) activity
  evidence_type: IBA
  original_reference_id: GO_REF:0000033
  qualifier: enables
  review:
    summary: >-
      Phylogenetic (IBA) annotation assigning long-chain 3-hydroxyacyl-CoA dehydrogenase
      (LCHAD, EC 1.1.1.211) activity to HADHA. This is the second core catalytic activity of
      the alpha subunit (step 3 of beta-oxidation, oxidation of (3S)-3-hydroxyacyl-CoA to
      3-oxoacyl-CoA using NAD+). The long-chain (rather than generic) term correctly reflects
      the C10-C16 substrate specificity of the enzyme.
    action: ACCEPT
    reason: >-
      Core molecular function of HADHA, supported by direct expression studies and
      long-chain substrate specificity data.
    supported_by:
    - reference_id: PMID:8135828
      supporting_text: "Expression of this cDNA in mammalian cells yielded a polypeptide with
        the long-chain enoyl-CoA hydratase and long-chain 3-hydroxyacyl-CoA dehydrogenase
        activities."
      full_text_unavailable: true
- term:
    id: GO:0016509
    label: long-chain (3S)-3-hydroxyacyl-CoA dehydrogenase (NAD+) activity
  evidence_type: IEA
  original_reference_id: GO_REF:0000120
  qualifier: enables
  review:
    summary: >-
      Electronic (IEA) annotation of long-chain 3-hydroxyacyl-CoA dehydrogenase activity
      based on RHEA/EC mapping. Consistent with the IBA and direct experimental evidence.
    action: ACCEPT
    reason: >-
      Redundant with experimental annotations but correctly captures the core dehydrogenase
      function with the appropriate long-chain specificity.
- term:
    id: GO:0016509
    label: long-chain (3S)-3-hydroxyacyl-CoA dehydrogenase (NAD+) activity
  evidence_type: EXP
  original_reference_id: PMID:1550553
  qualifier: enables
  review:
    summary: >-
      Experimental annotation. PMID:1550553 purified the long-chain-specific
      3-hydroxyacyl-CoA dehydrogenase from human liver mitochondrial membranes and showed
      maximum activity with C10-C16 substrates and inactivity with acetoacetyl-CoA, defining
      the long-chain specificity that distinguishes HADHA from the short-chain HADH.
    action: ACCEPT
    reason: >-
      Core molecular function with direct experimental support and explicit long-chain
      specificity.
    supported_by:
    - reference_id: PMID:1550553
      supporting_text: "gives maximum activity with 3-ketoacyl-CoA substrates of C10 to C16
        acyl-chain length but is inactive with acetoacetyl-CoA."
      full_text_unavailable: true
- term:
    id: GO:0016509
    label: long-chain (3S)-3-hydroxyacyl-CoA dehydrogenase (NAD+) activity
  evidence_type: EXP
  original_reference_id: PMID:8135828
  qualifier: enables
  review:
    summary: >-
      Experimental annotation citing the definitive subunit-assignment paper. Expression of
      the alpha-subunit cDNA yielded long-chain 3-hydroxyacyl-CoA dehydrogenase activity.
    action: ACCEPT
    reason: >-
      Core molecular function with strong direct evidence assigning the dehydrogenase to the
      alpha subunit.
    supported_by:
    - reference_id: PMID:8135828
      supporting_text: "Expression of this cDNA in mammalian cells yielded a polypeptide with
        the long-chain enoyl-CoA hydratase and long-chain 3-hydroxyacyl-CoA dehydrogenase
        activities."
      full_text_unavailable: true
- term:
    id: GO:0016509
    label: long-chain (3S)-3-hydroxyacyl-CoA dehydrogenase (NAD+) activity
  evidence_type: EXP
  original_reference_id: PMID:8163672
  qualifier: enables
  review:
    summary: >-
      Experimental annotation from the patient catalytic-heterogeneity study, which assayed
      the trifunctional protein's 3-hydroxyacyl-CoA dehydrogenase activity (with medium- to
      long-chain substrates) in fibroblasts from two LCHAD-deficiency patients.
    action: ACCEPT
    reason: >-
      Supports the dehydrogenase activity of the alpha subunit; the long-chain specific
      assignment is consistent with the gene's role in LCHAD deficiency.
    supported_by:
    - reference_id: PMID:8163672
      supporting_text: "We examined the enzyme protein and biosynthesis of human trifunctional
        protein harboring enoyl-CoA hydratase, 3-hydroxyacyl-CoA dehydrogenase, and
        3-ketoacyl-CoA thiolase activity in cultured skin fibroblasts from two patients with
        long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency."
      full_text_unavailable: true
- term:
    id: GO:0003857
    label: (3S)-3-hydroxyacyl-CoA dehydrogenase (NAD+) activity
  evidence_type: IEA
  original_reference_id: GO_REF:0000120
  qualifier: enables
  review:
    summary: >-
      Electronic annotation of the generic (chain-length-unspecified) 3-hydroxyacyl-CoA
      dehydrogenase activity. Correct in essence, but HADHA is long-chain specific, so the
      more specific term GO:0016509 better captures the molecular function.
    action: MODIFY
    reason: >-
      Generalize/replace with the long-chain-specific term GO:0016509, which is already
      present and is the appropriate level of specificity for HADHA (C10-C16 substrates).
    proposed_replacement_terms:
    - id: GO:0016509
      label: long-chain (3S)-3-hydroxyacyl-CoA dehydrogenase (NAD+) activity
- term:
    id: GO:0003857
    label: (3S)-3-hydroxyacyl-CoA dehydrogenase (NAD+) activity
  evidence_type: TAS
  original_reference_id: PMID:8135828
  qualifier: enables
  review:
    summary: >-
      TAS annotation of the generic 3-hydroxyacyl-CoA dehydrogenase activity citing
      PMID:8135828. The activity is correct for HADHA, but the long-chain specific term
      GO:0016509 is preferred given the demonstrated C10-C16 specificity.
    action: MODIFY
    reason: >-
      Generalize/replace with the long-chain-specific term GO:0016509.
    proposed_replacement_terms:
    - id: GO:0016509
      label: long-chain (3S)-3-hydroxyacyl-CoA dehydrogenase (NAD+) activity
    supported_by:
    - reference_id: PMID:8135828
      supporting_text: "Expression of this cDNA in mammalian cells yielded a polypeptide with
        the long-chain enoyl-CoA hydratase and long-chain 3-hydroxyacyl-CoA dehydrogenase
        activities."
      full_text_unavailable: true
# ============================================================================
# MOLECULAR FUNCTION - 3-HYDROXYACYL-CoA DEHYDRATASE (= hydratase, reverse direction)
# ============================================================================
- term:
    id: GO:0018812
    label: 3-hydroxyacyl-CoA dehydratase activity
  evidence_type: IEA
  original_reference_id: GO_REF:0000116
  qualifier: enables
  review:
    summary: >-
      RHEA-based electronic annotation. GO:0018812 describes the same chemical reaction as
      enoyl-CoA hydratase (RHEA:16105, 3-hydroxyacyl-CoA <=> 2-enoyl-CoA + H2O) written in the
      dehydration (reverse) direction. This is the alpha subunit's hydratase activity captured
      under a reverse-direction label.
    action: MODIFY
    reason: >-
      The conventional molecular-function term for this reaction in HADHA is enoyl-CoA
      hydratase activity (GO:0004300, EC 4.2.1.17, physiological hydration direction); the
      dehydratase label is the reverse framing of the same reaction.
    proposed_replacement_terms:
    - id: GO:0004300
      label: enoyl-CoA hydratase activity
- term:
    id: GO:0018812
    label: 3-hydroxyacyl-CoA dehydratase activity
  evidence_type: EXP
  original_reference_id: PMID:1550553
  qualifier: enables
  review:
    summary: >-
      Experimental annotation derived from the hydratase activity demonstrated in
      PMID:1550553. As above, this is the same reaction as enoyl-CoA hydratase written in the
      reverse direction.
    action: MODIFY
    reason: >-
      Replace with the conventional hydratase term GO:0004300 (EC 4.2.1.17).
    proposed_replacement_terms:
    - id: GO:0004300
      label: enoyl-CoA hydratase activity
    supported_by:
    - reference_id: PMID:1550553
      supporting_text: "the enzyme possesses 2-enoyl-CoA hydratase and 3-ketoacyl-CoA thiolase
        activities which cannot be separated from the dehydrogenase."
      full_text_unavailable: true
- term:
    id: GO:0018812
    label: 3-hydroxyacyl-CoA dehydratase activity
  evidence_type: EXP
  original_reference_id: PMID:8135828
  qualifier: enables
  review:
    summary: >-
      Experimental annotation from the subunit-assignment paper. The alpha subunit's hydratase
      activity is captured here under the reverse-direction dehydratase label.
    action: MODIFY
    reason: >-
      Replace with the conventional hydratase term GO:0004300 (EC 4.2.1.17).
    proposed_replacement_terms:
    - id: GO:0004300
      label: enoyl-CoA hydratase activity
    supported_by:
    - reference_id: PMID:8135828
      supporting_text: "Expression of this cDNA in mammalian cells yielded a polypeptide with
        the long-chain enoyl-CoA hydratase and long-chain 3-hydroxyacyl-CoA dehydrogenase
        activities."
      full_text_unavailable: true
# ============================================================================
# MOLECULAR FUNCTION - THIOLASE / ACETYLTRANSFERASE (BELONGS TO HADHB, NOT HADHA)
# ============================================================================
- term:
    id: GO:0003985
    label: acetyl-CoA C-acetyltransferase activity
  evidence_type: TAS
  original_reference_id: PMID:8135828
  qualifier: enables
  review:
    summary: >-
      This annotation assigns thiolase/acetyltransferase activity to HADHA, but PMID:8135828 -
      the very paper cited - showed that the 3-ketoacyl-CoA thiolase activity is carried by
      the BETA subunit (HADHB), not the alpha subunit. Expression of the alpha-subunit cDNA
      yielded only hydratase and dehydrogenase activities; only the beta-subunit cDNA yielded
      thiolase activity. The crystal structure (PMID:30850536) likewise places the
      3-ketothiolase active site in the beta subunit.
    action: REMOVE
    reason: >-
      Thiolase (acetyl-CoA C-acetyltransferase) activity belongs to HADHB, not HADHA, and is
      contradicted by the cited paper itself and by the structural data. This is an old PINC
      annotation that mis-attributed a beta-subunit activity to the alpha subunit.
    supported_by:
    - reference_id: PMID:8135828
      supporting_text: "Expression of this cDNA in mammalian cells yielded a polypeptide with
        the long-chain enoyl-CoA hydratase and long-chain 3-hydroxyacyl-CoA dehydrogenase
        activities."
      full_text_unavailable: true
    - reference_id: PMID:30850536
      supporting_text: "employing 2-enoyl-CoA hydratase (ECH), 3-hydroxyl-CoA dehydrogenase
        (HAD), and 3-ketothiolase (KT) activities consecutively."
      full_text_unavailable: true
# ============================================================================
# MOLECULAR FUNCTION - LONG-CHAIN FATTY ACYL-CoA HYDROLASE (WRONG PAPER)
# ============================================================================
- term:
    id: GO:0052816
    label: long-chain fatty acyl-CoA hydrolase activity
  evidence_type: IDA
  original_reference_id: PMID:22586271
  qualifier: enables
  review:
    summary: >-
      This IDA annotates HADHA with long-chain fatty acyl-CoA hydrolase (thioesterase)
      activity citing PMID:22586271. That paper (full text available) is a structural and
      functional study of the Hotdog-fold mitochondrial thioesterase Them5/Acot15 and its
      paralog Them4; it contains no mention of HADHA, the trifunctional protein, or P40939
      anywhere in the abstract or full text. The thioesterase activity it describes belongs to
      Them5, not HADHA. HADHA is a hydratase/dehydrogenase, not an acyl-CoA thioesterase.
    action: REMOVE
    reason: >-
      Wrong-paper attribution. The cited reference does not assay HADHA and does not support a
      thioesterase/hydrolase activity for this protein; the activity is that of Them5/Acot15.
    supported_by:
    - reference_id: PMID:22586271
      supporting_text: "Here, we present a structural and functional analysis of a new
        mammalian mitochondrial thioesterase, Them5."
# ============================================================================
# MOLECULAR FUNCTION - BROAD / PARENT TERMS (NON-CORE)
# ============================================================================
- term:
    id: GO:0003824
    label: catalytic activity
  evidence_type: IEA
  original_reference_id: GO_REF:0000002
  qualifier: enables
  review:
    summary: >-
      Root-level catalytic activity term from InterPro mapping. Correct but entirely
      uninformative.
    action: KEEP_AS_NON_CORE
    reason: >-
      Too general; the specific hydratase and dehydrogenase terms capture the molecular
      function.
- term:
    id: GO:0016491
    label: oxidoreductase activity
  evidence_type: IEA
  original_reference_id: GO_REF:0000002
  qualifier: enables
  review:
    summary: >-
      Broad oxidoreductase parent term from InterPro mapping, reflecting the dehydrogenase
      domain.
    action: KEEP_AS_NON_CORE
    reason: >-
      Correct but too general; GO:0016509 is the appropriate specific term.
- term:
    id: GO:0016616
    label: oxidoreductase activity, acting on the CH-OH group of donors, NAD or NADP
      as acceptor
  evidence_type: IEA
  original_reference_id: GO_REF:0000002
  qualifier: enables
  review:
    summary: >-
      Intermediate-specificity parent of the 3-hydroxyacyl-CoA dehydrogenase activity, from
      InterPro mapping.
    action: KEEP_AS_NON_CORE
    reason: >-
      Correct parent term; the specific child GO:0016509 should be used for core function.
- term:
    id: GO:0070403
    label: NAD+ binding
  evidence_type: IEA
  original_reference_id: GO_REF:0000002
  qualifier: enables
  review:
    summary: >-
      NAD+ binding from InterPro mapping, reflecting the NAD-binding Rossmann domain of the
      dehydrogenase. A cofactor-binding capability that supports but does not by itself
      describe the catalytic function.
    action: KEEP_AS_NON_CORE
    reason: >-
      Correct molecular characteristic (the dehydrogenase uses NAD+), but subsidiary to the
      catalytic dehydrogenase activity.
# ============================================================================
# MOLECULAR FUNCTION - PROTEIN BINDING (NON-CORE)
# ============================================================================
- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:20562859
  qualifier: enables
  review:
    summary: >-
      IPI from an autophagy-system network study (HADHA detected interacting with autophagy
      machinery, including GABARAP/ATG8 family members).
    action: KEEP_AS_NON_CORE
    reason: >-
      Generic protein binding is uninformative about molecular function; per curation
      guidelines it is not endorsed as core.
- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:28514442
  qualifier: enables
  review:
    summary: >-
      IPI based on interaction with HADHB (P55084), the obligate beta-subunit partner in the
      MTP complex.
    action: KEEP_AS_NON_CORE
    reason: >-
      Reflects obligate complex formation, which is better captured by the complex-membership
      annotation GO:0016507; generic protein binding is uninformative.
- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:29915090
  qualifier: enables
  review:
    summary: >-
      IPI from the cryo-EM structure study, reflecting HADHA-HADHB (P55084) interaction within
      the TFP tetramer.
    action: KEEP_AS_NON_CORE
    reason: >-
      Obligate complex interaction better represented by GO:0016507.
- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:30850536
  qualifier: enables
  review:
    summary: >-
      IPI from the crystal structure study, reflecting HADHA-HADHB (P55084) interaction in the
      alpha2-beta2 complex.
    action: KEEP_AS_NON_CORE
    reason: >-
      Obligate complex interaction better represented by GO:0016507.
- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:32814053
  qualifier: enables
  review:
    summary: >-
      IPI from a neurodegenerative-disease interactome mapping study.
    action: KEEP_AS_NON_CORE
    reason: >-
      Generic protein binding term; uninformative about molecular function.
- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:33961781
  qualifier: enables
  review:
    summary: >-
      IPI from a dual proteome-scale interactome study, again reflecting interaction with
      HADHB (P55084).
    action: KEEP_AS_NON_CORE
    reason: >-
      Obligate complex interaction better represented by GO:0016507.
- term:
    id: GO:0005515
    label: protein binding
  evidence_type: IPI
  original_reference_id: PMID:40205054
  qualifier: enables
  review:
    summary: >-
      IPI from a multimodal cell-maps study, reflecting interaction with HADHB (P55084).
    action: KEEP_AS_NON_CORE
    reason: >-
      Obligate complex interaction better represented by GO:0016507.
# ============================================================================
# BIOLOGICAL PROCESS - FATTY ACID BETA-OXIDATION (CORE)
# ============================================================================
- term:
    id: GO:0006635
    label: fatty acid beta-oxidation
  evidence_type: IBA
  original_reference_id: GO_REF:0000033
  qualifier: involved_in
  review:
    summary: >-
      Phylogenetic annotation placing HADHA in the fatty acid beta-oxidation pathway. The MTP
      complex, of which HADHA is the catalytic alpha subunit, performs the last three steps of
      long-chain fatty acid beta-oxidation.
    action: ACCEPT
    reason: >-
      Core biological process, supported by direct functional, genetic and structural data.
    supported_by:
    - reference_id: PMID:29915090
      supporting_text: "The mitochondrial trifunctional protein (TFP) catalyzes three reactions
        in the fatty acid beta-oxidation process."
      full_text_unavailable: true
- term:
    id: GO:0006635
    label: fatty acid beta-oxidation
  evidence_type: IEA
  original_reference_id: GO_REF:0000120
  qualifier: involved_in
  review:
    summary: >-
      Electronic annotation of fatty acid beta-oxidation, consistent with IBA and direct
      evidence.
    action: ACCEPT
    reason: >-
      Redundant but correctly captures the core biological process.
- term:
    id: GO:0006635
    label: fatty acid beta-oxidation
  evidence_type: IDA
  original_reference_id: PMID:29915090
  qualifier: involved_in
  review:
    summary: >-
      IDA (ComplexPortal/UniProt) from the cryo-EM structural characterization of the TFP
      complex in beta-oxidation.
    action: ACCEPT
    reason: >-
      Core biological process with direct structural evidence for the functional
      beta-oxidation complex.
    supported_by:
    - reference_id: PMID:29915090
      supporting_text: "The mitochondrial trifunctional protein (TFP) catalyzes three reactions
        in the fatty acid beta-oxidation process."
      full_text_unavailable: true
- term:
    id: GO:0006635
    label: fatty acid beta-oxidation
  evidence_type: IMP
  original_reference_id: PMID:31604922
  qualifier: involved_in
  review:
    summary: >-
      IMP from HADHA-deficient human iPSC-derived cardiomyocytes, which demonstrated that
      HADHA is required for fatty acid beta-oxidation (loss of HADHA produced reduced FAO and
      a metabolic/electrophysiological disease phenotype).
    action: ACCEPT
    reason: >-
      Core biological process supported by loss-of-function (mutant phenotype) evidence in a
      human cell model.
    supported_by:
    - reference_id: PMID:31604922
      supporting_text: "HADHA (tri-functional protein alpha), a monolysocardiolipin
        acyltransferase-like enzyme, is required for fatty acid beta-oxidation and cardiolipin
        remodeling, essential for functional mitochondria in human"
# ============================================================================
# BIOLOGICAL PROCESS - CARDIOLIPIN REMODELING (alpha-specific moonlighting)
# ============================================================================
- term:
    id: GO:0035965
    label: cardiolipin acyl-chain remodeling
  evidence_type: IDA
  original_reference_id: PMID:23152787
  qualifier: involved_in
  review:
    summary: >-
      IDA showing that the alpha subunit of TFP has monolysocardiolipin acyltransferase
      activity that remodels cardiolipin. Purified recombinant alphaTFP acylated MLCL to CL
      with linoleoyl-, oleoyl- and palmitoyl-CoA, and its expression increased linoleate/oleate
      incorporation into CL in cells, linking beta-oxidation to cardiolipin remodeling. This is
      an activity of the alpha subunit independent of the beta subunit.
    action: ACCEPT
    reason: >-
      Well-supported, alpha-subunit-specific moonlighting function in cardiolipin remodeling.
      Retained as a genuine (secondary) biological process for HADHA.
    supported_by:
    - reference_id: PMID:23152787
      supporting_text: "Purified human recombinant αTFP exhibited acyl-CoA acyltransferase
        activity in the acylation of MLCL to CL with linoleoyl-CoA, oleoyl-CoA and
        palmitoyl-CoA as substrates."
- term:
    id: GO:0035965
    label: cardiolipin acyl-chain remodeling
  evidence_type: IMP
  original_reference_id: PMID:31604922
  qualifier: involved_in
  review:
    summary: >-
      IMP from HADHA-deficient iPSC cardiomyocytes showing defective cardiolipin remodeling on
      loss of HADHA, corroborating the acyltransferase activity reported in PMID:23152787.
    action: ACCEPT
    reason: >-
      Mutant-phenotype evidence supporting HADHA's role in cardiolipin remodeling.
    supported_by:
    - reference_id: PMID:31604922
      supporting_text: "HADHA (tri-functional protein alpha), a monolysocardiolipin
        acyltransferase-like enzyme, is required for fatty acid beta-oxidation and cardiolipin
        remodeling, essential for functional mitochondria in human"
# ============================================================================
# BIOLOGICAL PROCESS - GENERAL FATTY ACID METABOLISM (NON-CORE)
# ============================================================================
- term:
    id: GO:0006631
    label: fatty acid metabolic process
  evidence_type: IEA
  original_reference_id: GO_REF:0000002
  qualifier: involved_in
  review:
    summary: >-
      Broad parent term of fatty acid beta-oxidation from InterPro mapping.
    action: KEEP_AS_NON_CORE
    reason: >-
      Correct but too general; GO:0006635 (fatty acid beta-oxidation) is the appropriate
      specific term.
# ============================================================================
# CELLULAR COMPONENT - MITOCHONDRIAL INNER MEMBRANE (CORE)
# ============================================================================
- term:
    id: GO:0005743
    label: mitochondrial inner membrane
  evidence_type: IEA
  original_reference_id: GO_REF:0000044
  qualifier: located_in
  review:
    summary: >-
      Electronic annotation of mitochondrial inner membrane localization based on UniProt
      subcellular location. The MTP complex is membrane-bound on the matrix face of the inner
      membrane.
    action: ACCEPT
    reason: >-
      Core cellular localization, supported by structural and direct evidence.
- term:
    id: GO:0005743
    label: mitochondrial inner membrane
  evidence_type: IDA
  original_reference_id: PMID:29915090
  qualifier: located_in
  review:
    summary: >-
      IDA (ComplexPortal) from the cryo-EM study, which showed that a concave surface of the
      TFP tetramer associates with the membrane.
    action: ACCEPT
    reason: >-
      Core cellular localization with direct structural evidence.
    supported_by:
    - reference_id: PMID:29915090
      supporting_text: "A concave surface of the TFP tetramer interacts with the detergent
        molecules in the structure, suggesting that this region is involved in associating
        with the membrane."
      full_text_unavailable: true
- term:
    id: GO:0005743
    label: mitochondrial inner membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-1482775
  qualifier: located_in
  review:
    summary: >-
      TAS from Reactome (MLCL acylation to CL by HADH at the inner membrane). Correct
      localization.
    action: ACCEPT
    reason: >-
      Correct inner membrane localization from pathway database curation.
- term:
    id: GO:0005743
    label: mitochondrial inner membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-77271
  qualifier: located_in
  review:
    summary: >-
      TAS from Reactome for a beta-oxidation reaction at the mitochondrial inner membrane.
    action: ACCEPT
    reason: >-
      Correct inner membrane localization for the beta-oxidation machinery.
- term:
    id: GO:0005743
    label: mitochondrial inner membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-77277
  qualifier: located_in
  review:
    summary: >-
      TAS from Reactome for the enoyl-CoA hydratase reaction (HADHA's own activity) at the
      inner membrane.
    action: ACCEPT
    reason: >-
      Correct inner membrane localization for the hydratase reaction carried by HADHA.
- term:
    id: GO:0005743
    label: mitochondrial inner membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-77283
  qualifier: located_in
  review:
    summary: >-
      TAS from Reactome for the 3-hydroxyacyl-CoA dehydrogenase reaction (HADHA's own
      activity) at the inner membrane.
    action: ACCEPT
    reason: >-
      Correct inner membrane localization for the dehydrogenase reaction carried by HADHA.
- term:
    id: GO:0005743
    label: mitochondrial inner membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-77301
  qualifier: located_in
  review:
    summary: >-
      TAS from Reactome for a C16 hydratase reaction at the inner membrane.
    action: ACCEPT
    reason: >-
      Correct inner membrane localization.
- term:
    id: GO:0005743
    label: mitochondrial inner membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-77303
  qualifier: located_in
  review:
    summary: >-
      TAS from Reactome for a C16 dehydrogenase reaction at the inner membrane.
    action: ACCEPT
    reason: >-
      Correct inner membrane localization.
- term:
    id: GO:0005743
    label: mitochondrial inner membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-77304
  qualifier: located_in
  review:
    summary: >-
      TAS from Reactome for a thiolase reaction at the inner membrane. Localization is correct
      for HADHA via complex membership even though the thiolase step is catalyzed by HADHB.
    action: ACCEPT
    reason: >-
      Correct inner membrane localization for the TFP complex.
- term:
    id: GO:0005743
    label: mitochondrial inner membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-77309
  qualifier: located_in
  review:
    summary: >-
      TAS from Reactome for a beta-oxidation reaction at the inner membrane.
    action: ACCEPT
    reason: >-
      Correct inner membrane localization.
- term:
    id: GO:0005743
    label: mitochondrial inner membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-77321
  qualifier: located_in
  review:
    summary: >-
      TAS from Reactome for a beta-oxidation reaction at the inner membrane.
    action: ACCEPT
    reason: >-
      Correct inner membrane localization.
- term:
    id: GO:0005743
    label: mitochondrial inner membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-77329
  qualifier: located_in
  review:
    summary: >-
      TAS from Reactome for a beta-oxidation reaction at the inner membrane.
    action: ACCEPT
    reason: >-
      Correct inner membrane localization.
- term:
    id: GO:0005743
    label: mitochondrial inner membrane
  evidence_type: TAS
  original_reference_id: Reactome:R-HSA-77340
  qualifier: located_in
  review:
    summary: >-
      TAS from Reactome for a beta-oxidation reaction at the inner membrane.
    action: ACCEPT
    reason: >-
      Correct inner membrane localization.
# ============================================================================
# CELLULAR COMPONENT - MITOCHONDRIAL COMPLEX (CORE)
# ============================================================================
- term:
    id: GO:0016507
    label: mitochondrial fatty acid beta-oxidation multienzyme complex
  evidence_type: IBA
  original_reference_id: GO_REF:0000033
  qualifier: part_of
  review:
    summary: >-
      Phylogenetic annotation placing HADHA as part of the mitochondrial TFP complex. HADHA is
      the alpha subunit of the alpha2-beta2 heterotetramer.
    action: ACCEPT
    reason: >-
      Core cellular component; HADHA is an obligate subunit of the TFP complex whose structure
      has been determined by cryo-EM and X-ray crystallography.
    supported_by:
    - reference_id: PMID:30850536
      supporting_text: "The biological unit of the protein is α2β2"
      full_text_unavailable: true
- term:
    id: GO:0016507
    label: mitochondrial fatty acid beta-oxidation multienzyme complex
  evidence_type: IEA
  original_reference_id: GO_REF:0000002
  qualifier: part_of
  review:
    summary: >-
      InterPro-based electronic annotation of TFP complex membership, consistent with IBA and
      structural evidence.
    action: ACCEPT
    reason: >-
      Redundant but correctly captures core complex membership.
- term:
    id: GO:0016507
    label: mitochondrial fatty acid beta-oxidation multienzyme complex
  evidence_type: IPI
  original_reference_id: PMID:29915090
  qualifier: part_of
  review:
    summary: >-
      IPI (ComplexPortal) from the cryo-EM structure directly demonstrating the alpha2-beta2
      tetrameric complex.
    action: ACCEPT
    reason: >-
      Core cellular component with direct structural evidence.
    supported_by:
    - reference_id: PMID:29915090
      supporting_text: "Here we report a 4.2-Å cryo-electron microscopy α2β2 tetrameric
        structure of the human TFP."
      full_text_unavailable: true
# ============================================================================
# CELLULAR COMPONENT - GENERAL MITOCHONDRION (NON-CORE)
# ============================================================================
- term:
    id: GO:0005739
    label: mitochondrion
  evidence_type: IEA
  original_reference_id: GO_REF:0000120
  qualifier: located_in
  review:
    summary: >-
      General mitochondrial localization from electronic mapping. Correct but less specific
      than the inner membrane annotation.
    action: KEEP_AS_NON_CORE
    reason: >-
      Correct but subsumed by the more specific GO:0005743 / GO:0016507.
- term:
    id: GO:0005739
    label: mitochondrion
  evidence_type: IDA
  original_reference_id: GO_REF:0000052
  qualifier: located_in
  review:
    summary: >-
      IDA from HPA immunofluorescence confirming mitochondrial localization.
    action: KEEP_AS_NON_CORE
    reason: >-
      Correct but less specific than inner membrane localization.
- term:
    id: GO:0005739
    label: mitochondrion
  evidence_type: EXP
  original_reference_id: PMID:29915090
  qualifier: located_in
  review:
    summary: >-
      Experimental mitochondrial localization associated with the structural study.
    action: KEEP_AS_NON_CORE
    reason: >-
      Correct but the more specific inner membrane / complex terms are preferred for core
      annotation.
- term:
    id: GO:0005739
    label: mitochondrion
  evidence_type: IDA
  original_reference_id: PMID:23152787
  qualifier: located_in
  review:
    summary: >-
      IDA mitochondrial localization from the cardiolipin-remodeling study.
    action: KEEP_AS_NON_CORE
    reason: >-
      Correct but less specific than inner membrane localization.
- term:
    id: GO:0005739
    label: mitochondrion
  evidence_type: HTP
  original_reference_id: PMID:34800366
  qualifier: located_in
  review:
    summary: >-
      HTP mitochondrial localization from a quantitative mitochondrial proteome study.
    action: KEEP_AS_NON_CORE
    reason: >-
      Correct mitochondrial localization from proteomics; less specific than inner membrane.
- term:
    id: GO:0005739
    label: mitochondrion
  evidence_type: HDA
  original_reference_id: PMID:20833797
  qualifier: located_in
  review:
    summary: >-
      HDA mitochondrial localization from a muscle mitochondrial phosphoproteome study.
    action: KEEP_AS_NON_CORE
    reason: >-
      Correct mitochondrial localization; less specific than inner membrane.
# ============================================================================
# CELLULAR COMPONENT - MITOCHONDRIAL NUCLEOID (NON-CORE)
# ============================================================================
- term:
    id: GO:0042645
    label: mitochondrial nucleoid
  evidence_type: IDA
  original_reference_id: PMID:18063578
  qualifier: located_in
  review:
    summary: >-
      IDA from a mitochondrial nucleoid proteomics study. HADHA was identified in native
      nucleoid preparations, but such metabolic proteins were not observed to cross-link to
      mtDNA, indicating peripheral association rather than a core nucleoid role.
    action: KEEP_AS_NON_CORE
    reason: >-
      Peripheral/co-purifying association, not a functional nucleoid component; the core
      localization is the inner membrane.
    supported_by:
    - reference_id: PMID:18063578
      supporting_text: "Several other metabolic proteins and chaperones identified in native
        nucleoids, including ATAD3, were not observed to cross-link to mtDNA."
references:
- id: GO_REF:0000002
  title: Gene Ontology annotation through association of InterPro records with GO
    terms
  findings: []
- id: GO_REF:0000033
  title: Annotation inferences using phylogenetic trees
  findings: []
- id: GO_REF:0000044
  title: Gene Ontology annotation based on UniProtKB/Swiss-Prot Subcellular Location
    vocabulary mapping, accompanied by conservative changes to GO terms applied by
    UniProt
  findings: []
- id: GO_REF:0000052
  title: Gene Ontology annotation based on curation of immunofluorescence data
  findings: []
- id: GO_REF:0000116
  title: Automatic Gene Ontology annotation based on Rhea mapping
  findings: []
- id: GO_REF:0000120
  title: Combined Automated Annotation using Multiple IEA Methods
  findings: []
- id: PMID:1550553
  title: Human liver long-chain 3-hydroxyacyl-coenzyme A dehydrogenase is a multifunctional
    membrane-bound beta-oxidation enzyme of mitochondria.
  findings:
  - statement: >-
      Purified the long-chain-specific multifunctional beta-oxidation enzyme from human
      liver mitochondrial membranes and showed it possesses hydratase, dehydrogenase and
      thiolase activities (maximal with C10-C16 substrates) that could not be separated. Did
      not assign activities to specific subunits; subunit assignment was later done by
      PMID:8135828.
  full_text_unavailable: true
  reference_review:
    relevance: HIGH
    correctness: VERIFIED
    review_notes: >-
      Abstract-only cache. Establishes the long-chain specificity and the multifunctional
      nature of the enzyme; supports the hydratase and dehydrogenase activities now assigned
      to the alpha subunit.
- id: PMID:8135828
  title: Structural analysis of cDNAs for subunits of human mitochondrial fatty acid
    beta-oxidation trifunctional protein.
  findings:
  - statement: >-
      Definitive subunit-activity assignment. Expression of the alpha-subunit cDNA yielded
      long-chain enoyl-CoA hydratase and long-chain 3-hydroxyacyl-CoA dehydrogenase
      activities; expression of the beta-subunit cDNA yielded only long-chain 3-ketoacyl-CoA
      thiolase activity.
  full_text_unavailable: true
  reference_review:
    relevance: HIGH
    correctness: VERIFIED
    review_notes: >-
      Abstract-only cache. The key reference establishing that HADHA carries hydratase +
      dehydrogenase and that thiolase is HADHB's; basis for REMOVE of thiolase from HADHA.
- id: PMID:8163672
  title: Mitochondrial trifunctional protein deficiency. Catalytic heterogeneity of
    the mutant enzyme in two patients.
  findings:
  - statement: >-
      Assayed the trifunctional protein's hydratase, dehydrogenase and thiolase activities in
      fibroblasts from two LCHAD-deficiency patients, demonstrating that the disease is caused
      by abnormality of the trifunctional protein.
  full_text_unavailable: true
  reference_review:
    relevance: HIGH
    correctness: VERIFIED
    review_notes: >-
      Abstract-only cache. Supports the dehydrogenase activity and disease relationship.
- id: PMID:18063578
  title: The layered structure of human mitochondrial DNA nucleoids.
  findings:
  - statement: >-
      Mitochondrial nucleoid proteomics; metabolic proteins identified in native nucleoids
      were not cross-linked to mtDNA, indicating peripheral association.
  reference_review:
    relevance: LOW
    correctness: VERIFIED
    review_notes: >-
      Supports a peripheral, non-core nucleoid association for HADHA.
- id: PMID:20562859
  title: Network organization of the human autophagy system.
  findings:
  - statement: >-
      Proteomic network of the human autophagy system; source of an HADHA protein-binding
      interaction annotation.
  reference_review:
    relevance: LOW
    correctness: VERIFIED
    review_notes: >-
      Supports a generic protein-binding annotation only.
- id: PMID:20833797
  title: Phosphoproteome analysis of functional mitochondria isolated from resting
    human muscle reveals extensive phosphorylation of inner membrane protein complexes
    and enzymes.
  findings:
  - statement: >-
      Muscle mitochondrial phosphoproteome study supporting mitochondrial localization of
      HADHA.
  reference_review:
    relevance: LOW
    correctness: VERIFIED
- id: PMID:22586271
  title: Acyl coenzyme A thioesterase Them5/Acot15 is involved in cardiolipin remodeling
    and fatty liver development.
  findings:
  - statement: >-
      Structural and functional analysis of the Hotdog-fold mitochondrial thioesterase
      Them5/Acot15 (and its paralog Them4). The paper does not study HADHA/TFP/P40939; the
      long-chain acyl-CoA thioesterase activity it reports belongs to Them5, not HADHA.
  reference_review:
    relevance: NONE
    correctness: WRONG_IDENTIFIER
    review_notes: >-
      Full text available and confirms this paper is entirely about Them5/Acot15 with no
      mention of HADHA. The GO:0052816 (long-chain fatty acyl-CoA hydrolase) IDA on HADHA
      citing this paper is a wrong-paper attribution; annotation marked for REMOVE.
- id: PMID:23152787
  title: Human trifunctional protein alpha links cardiolipin remodeling to beta-oxidation.
  findings:
  - statement: >-
      Purified recombinant alpha subunit of TFP (alphaTFP) exhibits monolysocardiolipin
      acyltransferase activity, acylating MLCL to cardiolipin with linoleoyl-, oleoyl- and
      palmitoyl-CoA, and its expression increases incorporation of these fatty acids into CL
      in cells. Establishes an alpha-subunit-specific cardiolipin remodeling activity
      independent of the beta subunit.
  reference_review:
    relevance: HIGH
    correctness: VERIFIED
    review_notes: >-
      Full text available; direct support for the cardiolipin acyl-chain remodeling
      (GO:0035965) annotation and the MLCL acyltransferase (EC 2.3.1.-) activity of HADHA.
- id: PMID:28514442
  title: Architecture of the human interactome defines protein communities and disease
    networks.
  findings:
  - statement: >-
      Large-scale interactome study; source of an HADHA-HADHB protein-binding interaction.
  reference_review:
    relevance: LOW
    correctness: VERIFIED
    review_notes: >-
      Supports generic protein binding (HADHA-HADHB complex), better captured by GO:0016507.
- id: PMID:29915090
  title: Cryo-EM structure of human mitochondrial trifunctional protein.
  findings:
  - statement: >-
      4.2-A cryo-EM structure of the human TFP alpha2-beta2 tetramer with a V-shaped
      architecture; a concave surface associates with the inner membrane. Supports complex
      membership, inner membrane localization and beta-oxidation function.
  full_text_unavailable: true
  reference_review:
    relevance: HIGH
    correctness: VERIFIED
    review_notes: >-
      Abstract-only cache but abstract directly supports the complex, localization and
      function claims.
- id: PMID:30850536
  title: Crystal structure of human mitochondrial trifunctional protein, a fatty acid
    β-oxidation metabolon.
  findings:
  - statement: >-
      3.6-A crystal structure of the human TFP alpha2-beta2 metabolon. Confirms that the ECH
      (hydratase) and HAD (dehydrogenase) active sites are in the alpha subunit and the
      3-ketothiolase (KT) site is in the beta subunit, with a substrate-channeling pathway
      along the inner membrane.
  full_text_unavailable: true
  reference_review:
    relevance: HIGH
    correctness: VERIFIED
    review_notes: >-
      Abstract-only cache; supports the alpha-subunit activity assignment (hydratase +
      dehydrogenase) and inner membrane association.
- id: PMID:31604922
  title: TFPa/HADHA is required for fatty acid beta-oxidation and cardiolipin re-modeling
    in human cardiomyocytes.
  findings:
  - statement: >-
      HADHA-deficient human iPSC-derived cardiomyocytes show reduced fatty acid beta-oxidation
      and defective cardiolipin remodeling, demonstrating that HADHA is required for both
      processes. Provides loss-of-function (IMP) support for GO:0006635 and GO:0035965.
  reference_review:
    relevance: HIGH
    correctness: VERIFIED
    review_notes: >-
      Full text available; directly supports the beta-oxidation and cardiolipin remodeling
      mutant-phenotype annotations.
- id: PMID:32814053
  title: Interactome Mapping Provides a Network of Neurodegenerative Disease Proteins
    and Uncovers Widespread Protein Aggregation in Affected Brains.
  findings:
  - statement: >-
      Neurodegenerative-disease interactome mapping; source of an HADHA protein-binding
      interaction annotation.
  reference_review:
    relevance: LOW
    correctness: VERIFIED
- id: PMID:33961781
  title: Dual proteome-scale networks reveal cell-specific remodeling of the human
    interactome.
  findings:
  - statement: >-
      Dual proteome-scale interactome study; source of an HADHA-HADHB protein-binding
      interaction annotation.
  reference_review:
    relevance: LOW
    correctness: VERIFIED
- id: PMID:34800366
  title: Quantitative high-confidence human mitochondrial proteome and its dynamics
    in cellular context.
  findings:
  - statement: >-
      High-confidence mitochondrial proteome study confirming mitochondrial localization of
      HADHA.
  reference_review:
    relevance: LOW
    correctness: VERIFIED
- id: PMID:40205054
  title: Multimodal cell maps as a foundation for structural and functional genomics.
  findings:
  - statement: >-
      Multimodal cell-mapping study integrating interaction and imaging data; source of an
      HADHA-HADHB protein-binding interaction annotation.
  reference_review:
    relevance: LOW
    correctness: VERIFIED
- id: PMID:8651282
  title: 'Molecular characterization of mitochondrial trifunctional protein deficiency:
    formation of the enzyme complex is important for stabilization of both alpha- and
    beta-subunits.'
  findings:
  - statement: >-
      Cited in UniProt for catalytic activity of HADHA; shows that formation of the
      alpha-beta enzyme complex is important for stabilization of both subunits.
  full_text_unavailable: true
  reference_review:
    relevance: MEDIUM
    correctness: UNVERIFIED
    review_notes: >-
      Not present in the publications cache; not used as a supporting_text source. Listed for
      completeness as a UniProt-cited catalytic-activity reference.
- id: Reactome:R-HSA-1482775
  title: MLCL is acylated to CL by HADH (IM)
  findings:
  - statement: >-
      Reactome reaction: acylation of monolysocardiolipin (MLCL) to cardiolipin (CL) at the
      mitochondrial inner membrane, consistent with the HADHA MLCL acyltransferase activity.
- id: Reactome:R-HSA-77271
  title: 3-Oxotetradecanoyl-CoA+CoA-SH<=>Lauroyl-CoA
  findings:
  - statement: >-
      Thiolase step of beta-oxidation (C14) at the inner membrane; part of the TFP-catalyzed
      pathway (thiolysis is HADHB's activity, but the complex including HADHA acts here).
- id: Reactome:R-HSA-77277
  title: trans-Tetradec-2-enoyl-CoA+H2O<=>(S)-3-Hydroxytetradecanoyl-CoA
  findings:
  - statement: >-
      Enoyl-CoA hydratase reaction (C14) at the inner membrane - this is HADHA's own hydratase
      activity.
- id: Reactome:R-HSA-77283
  title: (S)-3-Hydroxytetradecanoyl-CoA+NAD<=>3-Oxotetradecanoyl-CoA+NADH+H
  findings:
  - statement: >-
      3-Hydroxyacyl-CoA dehydrogenase reaction (C14) at the inner membrane - this is HADHA's
      own dehydrogenase activity.
- id: Reactome:R-HSA-77301
  title: trans-Hexadec-2-enoyl-CoA+H2O<=>(S)-3-Hydroxyhexadecanoyl-CoA
  findings:
  - statement: >-
      Enoyl-CoA hydratase reaction (C16) - HADHA's hydratase activity.
- id: Reactome:R-HSA-77303
  title: (S)-3-Hydroxyhexadecanoyl-CoA+NAD<=>3-Oxopalmitoyl-CoA+NADH+H
  findings:
  - statement: >-
      3-Hydroxyacyl-CoA dehydrogenase reaction (C16) - HADHA's dehydrogenase activity.
- id: Reactome:R-HSA-77304
  title: 3-Oxopalmitoyl-CoA+CoA-SH<=>myristoyl-CoA
  findings:
  - statement: >-
      Thiolase step of beta-oxidation (C16) - HADHB's activity within the TFP complex.
- id: Reactome:R-HSA-77309
  title: 3-Oxododecanoyl-CoA+CoA-SH<=>Decanoyl-CoA
  findings:
  - statement: >-
      Thiolase step of beta-oxidation (C12) within the TFP complex.
- id: Reactome:R-HSA-77321
  title: 3-Oxohexanoyl-CoA+CoA-SH<=>Butanoyl-CoA
  findings:
  - statement: >-
      Thiolase step (C6) within the TFP pathway; at the short-chain edge of/below the
      complex's preferred long-chain range.
- id: Reactome:R-HSA-77329
  title: 3-Oxooctanoyl-CoA+CoA-SH<=>Hexanoyl-CoA
  findings:
  - statement: >-
      Thiolase step (C8) within the TFP pathway; medium-chain, at the edge of the complex's
      preferred long-chain range.
- id: Reactome:R-HSA-77340
  title: 3-Oxodecanoyl-CoA+CoA-SH<=>Octanoyl-CoA
  findings:
  - statement: >-
      Thiolase step (C10) within the TFP pathway; within the long-chain range.
core_functions:
- description: >-
    Long-chain enoyl-CoA hydratase (LCEH, EC 4.2.1.17): the second step of each
    mitochondrial long-chain fatty acid beta-oxidation cycle, catalyzing hydration of a
    (2E)-enoyl-CoA to (3S)-3-hydroxyacyl-CoA. This activity resides in the alpha subunit of
    the membrane-bound trifunctional protein and is specific for long-chain substrates.
  supported_by:
  - reference_id: PMID:8135828
    supporting_text: "Expression of this cDNA in mammalian cells yielded a polypeptide with
      the long-chain enoyl-CoA hydratase and long-chain 3-hydroxyacyl-CoA dehydrogenase
      activities."
    full_text_unavailable: true
  - reference_id: PMID:30850536
    supporting_text: "employing 2-enoyl-CoA hydratase (ECH), 3-hydroxyl-CoA dehydrogenase
      (HAD), and 3-ketothiolase (KT) activities consecutively."
    full_text_unavailable: true
  molecular_function:
    id: GO:0004300
    label: enoyl-CoA hydratase activity
  directly_involved_in:
  - id: GO:0006635
    label: fatty acid beta-oxidation
  locations:
  - id: GO:0005743
    label: mitochondrial inner membrane
  in_complex:
    id: GO:0016507
    label: mitochondrial fatty acid beta-oxidation multienzyme complex
- description: >-
    Long-chain 3-hydroxyacyl-CoA dehydrogenase (LCHAD, EC 1.1.1.211): the third step of
    each long-chain fatty acid beta-oxidation cycle, oxidizing (3S)-3-hydroxyacyl-CoA to
    3-oxoacyl-CoA with reduction of NAD+ to NADH. This NAD+-dependent activity resides in the
    alpha subunit of the trifunctional protein and is specific for long-chain (C10-C16)
    substrates; its deficiency causes LCHAD deficiency.
  supported_by:
  - reference_id: PMID:8135828
    supporting_text: "Expression of this cDNA in mammalian cells yielded a polypeptide with
      the long-chain enoyl-CoA hydratase and long-chain 3-hydroxyacyl-CoA dehydrogenase
      activities."
    full_text_unavailable: true
  - reference_id: PMID:1550553
    supporting_text: "gives maximum activity with 3-ketoacyl-CoA substrates of C10 to C16
      acyl-chain length but is inactive with acetoacetyl-CoA."
    full_text_unavailable: true
  molecular_function:
    id: GO:0016509
    label: long-chain (3S)-3-hydroxyacyl-CoA dehydrogenase (NAD+) activity
  directly_involved_in:
  - id: GO:0006635
    label: fatty acid beta-oxidation
  locations:
  - id: GO:0005743
    label: mitochondrial inner membrane
  in_complex:
    id: GO:0016507
    label: mitochondrial fatty acid beta-oxidation multienzyme complex
- description: >-
    Monolysocardiolipin (MLCL) acyltransferase (EC 2.3.1.-): an alpha-subunit-specific
    moonlighting activity that transfers an acyl group (linoleoyl, oleoyl or palmitoyl) from
    acyl-CoA onto monolysocardiolipin, reacylating it to mature cardiolipin. This activity is
    independent of the beta subunit and couples long-chain fatty acid beta-oxidation to
    cardiolipin acyl-chain remodeling, which is required for functional mitochondria. GO has no
    cardiolipin-specific molecular-function term, so the general acyltransferase activity term is
    used here pending a dedicated MLCL acyltransferase term.
  supported_by:
  - reference_id: PMID:23152787
    supporting_text: "Purified human recombinant αTFP exhibited acyl-CoA acyltransferase
      activity in the acylation of MLCL to CL with linoleoyl-CoA, oleoyl-CoA and
      palmitoyl-CoA as substrates."
  - reference_id: PMID:31604922
    supporting_text: "HADHA (tri-functional protein alpha), a monolysocardiolipin
      acyltransferase-like enzyme, is required for fatty acid beta-oxidation and cardiolipin
      remodeling, essential for functional mitochondria in human"
  molecular_function:
    id: GO:0016747
    label: acyltransferase activity, transferring groups other than amino-acyl groups
  directly_involved_in:
  - id: GO:0035965
    label: cardiolipin acyl-chain remodeling
  locations:
  - id: GO:0005743
    label: mitochondrial inner membrane
proposed_new_terms: []
suggested_questions:
- question: >-
    Is the monolysocardiolipin acyltransferase / cardiolipin remodeling activity of the alpha
    subunit a genuine independent physiological function in vivo, or a secondary moonlighting
    activity? Should it be modeled with a specific acyltransferase molecular-function term
    (EC 2.3.1.-) in addition to GO:0035965?
- question: >-
    Should the generic dehydrogenase (GO:0003857) and dehydratase (GO:0018812) annotations on
    HADHA be superseded by the long-chain-specific terms (GO:0016509, GO:0004300) to better
    reflect the demonstrated C10-C16 substrate specificity?
suggested_experiments:
- description: >-
    Reconstitute recombinant HADHA alone and as the alpha2-beta2 complex with HADHB and assay
    enoyl-CoA hydratase and 3-hydroxyacyl-CoA dehydrogenase activities across a chain-length
    series (C4-C20) to quantitatively define substrate specificity.
  hypothesis: >-
    HADHA exhibits hydratase and dehydrogenase activity restricted to long-chain (>=C10)
    acyl-CoA substrates, distinguishing it from the short-chain enzymes ECHS1 and HADH.
- description: >-
    Test whether the MLCL acyltransferase / cardiolipin remodeling activity of HADHA can be
    genetically separated from its hydratase/dehydrogenase activities using active-site point
    mutants in HADHA-deficient cardiomyocytes.
  hypothesis: >-
    The cardiolipin acyltransferase activity is mechanistically distinct from the
    beta-oxidation catalytic sites and can be uncoupled, indicating a bona fide moonlighting
    function.