ldhd encodes the mitochondrial FAD-dependent D-lactate dehydrogenase, a Mn2+-dependent flavoprotein of the inner mitochondrial membrane that oxidizes D-lactate and other (2R)-2-hydroxycarboxylates with small hydrophobic C2 substituents to the corresponding 2-oxoacids, passing the electrons to cytochrome c. It is strictly stereoselective (no activity on L-lactate) and clears D-lactate generated endogenously via the methylglyoxal pathway or taken up from gut bacteria and diet. Zebrafish ldhd mutants are viable and fertile but accumulate D-lactate, the biochemical hallmark of human D-lactate dehydrogenase deficiency.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:1903457 lactate catabolic process | IBA GO_REF:0000033 | ACCEPT | Summary: Lactate catabolic process: Ldhd performs the committed step of D-lactate breakdown, oxidizing D-lactate to pyruvate. The IBA descends from PTN000202298 (yeast DLD1, human LDHD) and is independently confirmed in zebrafish. Reason: The gene product itself catalyses the catabolic reaction (participation, not merely necessity), the loss-of-function metabolite phenotype is specific to the D-enantiomer, and the rescue by human LDHD shows the same activity is conserved; this is the core process for this enzyme. Supporting Evidence: PMID:30931947 metabolic analysis revealed elevated levels of D-lactate, but not L-lactate, in ldhd−/− larvae compared to wildtype PMID:30931947 ldhd−/− uninjected embryos showed a significant D-lactate increase that was restored to baseline levels by mRNA microinjection of the wildtype LDHD sequence file:DANRE/ldhd/ldhd-uniprot.txt prevents the toxic accumulation of D-lactate in the organism |
| GO:0003824 catalytic activity | IEA GO_REF:0000002 | MODIFY | Summary: Catalytic activity (InterPro FAD-binding oxidoreductase mapping) is correct but uninformative: the enzyme is an FAD-dependent D-lactate/(2R)-2-hydroxycarboxylate dehydrogenase with experimental support in zebrafish [PMID:30931947] and orthologs [PMID:37863926]. Reason: The root MF term adds nothing beyond the specific Rhea-mapped and experimental rows; replace with the specific FAD-dependent dehydrogenase terms already carried by EXP/ISS/IEA rows. Proposed replacements: D-lactate dehydrogenase (FAD) activity (2R)-2-hydroxycarboxylate dehydrogenase (FAD) activity Supporting Evidence: file:DANRE/ldhd/ldhd-uniprot.txt Reaction=(R)-lactate + FAD + H(+) = FADH2 + pyruvate file:DANRE/ldhd/ldhd-uniprot.txt Reaction=a (2R)-2-hydroxycarboxylate + FAD + H(+) = a 2-oxocarboxylate PMID:30931947 establishing that LDHD is stereospecific for D-lactate only |
| GO:0005739 mitochondrion | IEA GO_REF:0000044 | ACCEPT | Summary: Mitochondrion: UniProt SubCell-mapped IEA; the zebrafish protein is annotated mitochondrial by similarity, and the mammalian orthologs are inner-mitochondrial-membrane flavoproteins that feed electrons to cytochrome c [PMID:37863926; PMID:38413804]. Reason: Mitochondrial localization is the defining property of this FAD-linked D-lactate dehydrogenase lineage (yeast DLD1 to mammalian LDHD); there is no zebrafish-specific evidence of divergence and the D-lactate metabolite phenotype in ldhd mutants is consistent with the conserved mitochondrial role. Accepted as the site of the core activity. Supporting Evidence: file:DANRE/ldhd/ldhd-uniprot.txt SUBCELLULAR LOCATION: Mitochondrion PMID:37863926 FAD-dependent D-LDHs were also identified in mammals including human and mouse, which are localized in the mitochondrial inner membrane PMID:38413804 we could confirm direct electron transfer from d-lactate to cytochrome c by monitoring its haem absorbance |
| GO:0050660 flavin adenine dinucleotide binding | IEA GO_REF:0000002 | KEEP AS NON CORE | Summary: Flavin adenine dinucleotide binding: the enzyme carries a PCMH-type FAD-binding domain and uses FAD as the hydride acceptor; mouse LDHD crystal structures are FAD-bound and the reaction is defined with FAD as cosubstrate [PMID:37863926]. Reason: FAD binding is real and required (the catalytic reaction is written with FAD), but it is a cofactor-binding property subsumed by the FAD-dependent dehydrogenase activity term; kept as non-core so the core function remains the dehydrogenase activity. Supporting Evidence: file:DANRE/ldhd/ldhd-uniprot.txt FAD-binding PCMH-type file:DANRE/ldhd/ldhd-uniprot.txt Reaction=(R)-lactate + FAD + H(+) = FADH2 + pyruvate PMID:37863926 crystal structures of mLDHD in FAD-bound |
| GO:0071949 FAD binding | IEA GO_REF:0000002 | KEEP AS NON CORE | Summary: FAD binding: the enzyme carries a PCMH-type FAD-binding domain and uses FAD as the hydride acceptor; mouse LDHD crystal structures are FAD-bound and the reaction is defined with FAD as cosubstrate [PMID:37863926]. Reason: FAD binding is real and required (the catalytic reaction is written with FAD), but it is a cofactor-binding property subsumed by the FAD-dependent dehydrogenase activity term; kept as non-core so the core function remains the dehydrogenase activity. Supporting Evidence: file:DANRE/ldhd/ldhd-uniprot.txt FAD-binding PCMH-type file:DANRE/ldhd/ldhd-uniprot.txt Reaction=(R)-lactate + FAD + H(+) = FADH2 + pyruvate PMID:37863926 crystal structures of mLDHD in FAD-bound |
| GO:0005739 mitochondrion | ISS GO_REF:0000024 | ACCEPT | Summary: Mitochondrion: ISS from mouse Ldhd (Q7TNG8); the zebrafish protein is annotated mitochondrial by similarity, and the mammalian orthologs are inner-mitochondrial-membrane flavoproteins that feed electrons to cytochrome c [PMID:37863926; PMID:38413804]. Reason: Mitochondrial localization is the defining property of this FAD-linked D-lactate dehydrogenase lineage (yeast DLD1 to mammalian LDHD); there is no zebrafish-specific evidence of divergence and the D-lactate metabolite phenotype in ldhd mutants is consistent with the conserved mitochondrial role. Accepted as the site of the core activity. Supporting Evidence: file:DANRE/ldhd/ldhd-uniprot.txt SUBCELLULAR LOCATION: Mitochondrion PMID:37863926 FAD-dependent D-LDHs were also identified in mammals including human and mouse, which are localized in the mitochondrial inner membrane PMID:38413804 we could confirm direct electron transfer from d-lactate to cytochrome c by monitoring its haem absorbance |
| GO:0140170 D-lactate dehydrogenase (FAD) activity | EXP PMID:30931947 Identification of human D lactate dehydrogenase deficiency. | ACCEPT | Summary: D-lactate dehydrogenase (FAD) activity (the zebrafish EXP row from ldhd mutants and human mRNA rescue): FAD-dependent oxidation of D-lactate to pyruvate. Zebrafish ldhd mutants accumulate D-lactate specifically and are rescued by human LDHD [PMID:30931947]; purified mouse and human LDHD oxidize D-lactate with FAD/Mn2+ and transfer electrons to cytochrome c [PMID:37863926; PMID:38413804]. Reason: This is the core molecular function. The zebrafish in vivo readout (D- but not L-lactate elevation, rescued by wild-type but not patient-variant human LDHD) demonstrates the enzyme's stereospecific activity in the fish, and the reaction chemistry (FAD as acceptor, Mn2+ cofactor) is established on the mammalian orthologs. Supporting Evidence: PMID:30931947 metabolic analysis revealed elevated levels of D-lactate, but not L-lactate, in ldhd−/− larvae compared to wildtype PMID:30931947 establishing that LDHD is stereospecific for D-lactate only file:DANRE/ldhd/ldhd-uniprot.txt Reaction=(R)-lactate + FAD + H(+) = FADH2 + pyruvate PMID:37863926 mLDHD is an Mn2+-dependent general dehydrogenase for a broad range of D-2-hydroxyacids with small to moderate-size hydrophobic moieties at the C2 atom |
| GO:0140174 (2R)-2-hydroxycarboxylate dehydrogenase (FAD) activity | EXP PMID:30931947 Identification of human D lactate dehydrogenase deficiency. | ACCEPT | Summary: (2R)-2-hydroxycarboxylate dehydrogenase (FAD) activity (zebrafish EXP row): the broader substrate class. Mouse LDHD acts on D-2-hydroxybutanoate through D-2-hydroxyoctanoate and branched D-2-hydroxyacids with hydrophobic C2 groups, but not on hydrophilic ones or L-isomers [PMID:37863926]; human LDHD reduces cytochrome c with D-2-hydroxybutyrate as well [PMID:38413804]. Reason: D-lactate is one member of the substrate class this enzyme handles; the generic (2R)-2-hydroxycarboxylate term captures the true substrate scope shown biochemically on the mammalian orthologs, and the zebrafish enzyme is the ortholog whose D-lactate activity is confirmed in vivo. Accepted alongside the D-lactate-specific term. Supporting Evidence: file:DANRE/ldhd/ldhd-uniprot.txt Reaction=a (2R)-2-hydroxycarboxylate + FAD + H(+) = a 2-oxocarboxylate PMID:37863926 mLDHD is an Mn2+-dependent general dehydrogenase for a broad range of D-2-hydroxyacids with small to moderate-size hydrophobic moieties at the C2 atom PMID:38413804 we identified d-2-hydroxybutyrate as another LDHD substrate that can be used to reduce cytochrome c |
| GO:0140170 D-lactate dehydrogenase (FAD) activity | ISS GO_REF:0000024 | ACCEPT | Summary: D-lactate dehydrogenase (FAD) activity (ISS from mouse Ldhd): FAD-dependent oxidation of D-lactate to pyruvate. Zebrafish ldhd mutants accumulate D-lactate specifically and are rescued by human LDHD [PMID:30931947]; purified mouse and human LDHD oxidize D-lactate with FAD/Mn2+ and transfer electrons to cytochrome c [PMID:37863926; PMID:38413804]. Reason: This is the core molecular function. The zebrafish in vivo readout (D- but not L-lactate elevation, rescued by wild-type but not patient-variant human LDHD) demonstrates the enzyme's stereospecific activity in the fish, and the reaction chemistry (FAD as acceptor, Mn2+ cofactor) is established on the mammalian orthologs. Supporting Evidence: PMID:30931947 metabolic analysis revealed elevated levels of D-lactate, but not L-lactate, in ldhd−/− larvae compared to wildtype PMID:30931947 establishing that LDHD is stereospecific for D-lactate only file:DANRE/ldhd/ldhd-uniprot.txt Reaction=(R)-lactate + FAD + H(+) = FADH2 + pyruvate PMID:37863926 mLDHD is an Mn2+-dependent general dehydrogenase for a broad range of D-2-hydroxyacids with small to moderate-size hydrophobic moieties at the C2 atom |
| GO:0140174 (2R)-2-hydroxycarboxylate dehydrogenase (FAD) activity | ISS GO_REF:0000024 | ACCEPT | Summary: (2R)-2-hydroxycarboxylate dehydrogenase (FAD) activity (ISS from mouse Ldhd): the broader substrate class. Mouse LDHD acts on D-2-hydroxybutanoate through D-2-hydroxyoctanoate and branched D-2-hydroxyacids with hydrophobic C2 groups, but not on hydrophilic ones or L-isomers [PMID:37863926]; human LDHD reduces cytochrome c with D-2-hydroxybutyrate as well [PMID:38413804]. Reason: D-lactate is one member of the substrate class this enzyme handles; the generic (2R)-2-hydroxycarboxylate term captures the true substrate scope shown biochemically on the mammalian orthologs, and the zebrafish enzyme is the ortholog whose D-lactate activity is confirmed in vivo. Accepted alongside the D-lactate-specific term. Supporting Evidence: file:DANRE/ldhd/ldhd-uniprot.txt Reaction=a (2R)-2-hydroxycarboxylate + FAD + H(+) = a 2-oxocarboxylate PMID:37863926 mLDHD is an Mn2+-dependent general dehydrogenase for a broad range of D-2-hydroxyacids with small to moderate-size hydrophobic moieties at the C2 atom PMID:38413804 we identified d-2-hydroxybutyrate as another LDHD substrate that can be used to reduce cytochrome c |
| GO:1903457 lactate catabolic process | IMP PMID:30931947 Identification of human D lactate dehydrogenase deficiency. | ACCEPT | Summary: Lactate catabolic process: Ldhd performs the committed step of D-lactate breakdown, oxidizing D-lactate to pyruvate. The IMP is from the ldhd sa15623 splice-site allele: maternal-zygotic mutant larvae accumulate D-lactate but not L-lactate, and human wild-type LDHD mRNA (not patient variants) rescues the accumulation [PMID:30931947]. Reason: The gene product itself catalyses the catabolic reaction (participation, not merely necessity), the loss-of-function metabolite phenotype is specific to the D-enantiomer, and the rescue by human LDHD shows the same activity is conserved; this is the core process for this enzyme. Supporting Evidence: PMID:30931947 metabolic analysis revealed elevated levels of D-lactate, but not L-lactate, in ldhd−/− larvae compared to wildtype PMID:30931947 ldhd−/− uninjected embryos showed a significant D-lactate increase that was restored to baseline levels by mRNA microinjection of the wildtype LDHD sequence file:DANRE/ldhd/ldhd-uniprot.txt prevents the toxic accumulation of D-lactate in the organism |
| GO:0004458 D-lactate dehydrogenase (cytochrome) activity | IBA GO_REF:0000033 | ACCEPT | Summary: D-lactate dehydrogenase (cytochrome) activity (IBA from PTN000202298: yeast DLD1, Arabidopsis DLD): oxidation of D-lactate with cytochrome c as electron acceptor. Purified human LDHD transfers electrons directly from D-lactate to cytochrome c (kcat 103 min-1, KM 112 uM) and LDHD-knockout cells lose the D-lactate-driven membrane potential response [PMID:38413804]. Reason: The PAINT node placement is sound: the cytochrome c-coupled D-lactate dehydrogenase activity of yeast DLD1 was inherited by the vertebrate LDHD clade, as shown directly for human LDHD in 2024. This is the physiological form of the FAD-dependent activity (FADH2 is reoxidized by cytochrome c), so it complements rather than duplicates GO:0140170. The zebrafish enzyme has not been assayed with cytochrome c, but its in vivo D-lactate clearance and conserved FAD/cytochrome-binding architecture give no reason to expect divergence. Supporting Evidence: PMID:38413804 we could confirm direct electron transfer from d-lactate to cytochrome c by monitoring its haem absorbance PMID:38413804 their response to d-lactate was blunted PMID:30931947 metabolic analysis revealed elevated levels of D-lactate, but not L-lactate, in ldhd−/− larvae compared to wildtype file:DANRE/ldhd/ldhd-deep-research-falcon.md A 2024 study provided direct evidence that **human LDHD transfers electrons from D-lactate to cytochrome c**, thereby supporting mitochondrial membrane polarization. |
| GO:0005739 mitochondrion | IBA GO_REF:0000033 | ACCEPT | Summary: Mitochondrion: IBA from PTN000202576 (yeast DLD1, mouse/rat Ldhd, Arabidopsis DLD); the zebrafish protein is annotated mitochondrial by similarity, and the mammalian orthologs are inner-mitochondrial-membrane flavoproteins that feed electrons to cytochrome c [PMID:37863926; PMID:38413804]. Reason: Mitochondrial localization is the defining property of this FAD-linked D-lactate dehydrogenase lineage (yeast DLD1 to mammalian LDHD); there is no zebrafish-specific evidence of divergence and the D-lactate metabolite phenotype in ldhd mutants is consistent with the conserved mitochondrial role. Accepted as the site of the core activity. Supporting Evidence: file:DANRE/ldhd/ldhd-uniprot.txt SUBCELLULAR LOCATION: Mitochondrion PMID:37863926 FAD-dependent D-LDHs were also identified in mammals including human and mouse, which are localized in the mitochondrial inner membrane PMID:38413804 we could confirm direct electron transfer from d-lactate to cytochrome c by monitoring its haem absorbance |
| GO:0008720 D-lactate dehydrogenase (NAD+) activity | IBA GO_REF:0000033 | MODIFY | Summary: D-lactate dehydrogenase (NAD+) activity (IBA from PTN000202298 with Arabidopsis AT5G06580 as the only experimental donor, IMP PMID:22155004 via QuickGO). The claim is NAD+-dependent D-lactate oxidation. LDHD is an FAD-linked PCMH-type flavoprotein, not an NAD-dependent 2-hydroxyacid dehydrogenase; mouse LDHD is assayed with FAD/Mn2+ and human LDHD with cytochrome c as acceptor [PMID:37863926; PMID:38413804]. Reason: The NAD+-dependent term describes a different enzyme family (bacterial D-isomer-specific 2-hydroxyacid dehydrogenases). The donor annotation on Arabidopsis DLD is a plant-selection-marker IMP that does not establish NAD+ as acceptor, and the vertebrate enzymes are characterised with FAD and cytochrome c, so the activity should be captured by the FAD-dependent term GO:0140170 (already on the gene) rather than the NAD+ term. Propagation Review Root cause: SOURCE WEAK OR INFERRED Failure modes: SOURCE EVIDENCE WEAK ROLE CONFLATION Sources checked: PANTHER:PTN000202298 SOURCE WEAK OR INFERRED Node carries the NAD+ term from a single plant IMP (AT5G06580); the family is FAD-linked and vertebrate members are assayed with FAD and cytochrome c, so the acceptor in the term does not match the enzyme chemistry at this node. AGI_LocusCode:AT5G06580 SOURCE WEAK OR INFERRED Arabidopsis DLD GO:0008720 IMP (PMID:22155004) rests on D-lactate tolerance as a selection marker, not on NAD+-dependent kinetics; the same enzyme is described elsewhere as FAD/cytochrome c-linked. Proposed replacements: D-lactate dehydrogenase (FAD) activity Supporting Evidence: PMID:37863926 The NAD-dependent D-LDHs were found in some bacteria and belong to the D-isomer specific 2-hydroxyacid dehydrogenase superfamily PMID:37863926 mLDHD is an Mn2+-dependent general dehydrogenase for a broad range of D-2-hydroxyacids with small to moderate-size hydrophobic moieties at the C2 atom file:DANRE/ldhd/ldhd-uniprot.txt Reaction=(R)-lactate + FAD + H(+) = FADH2 + pyruvate PMID:22155004 plants overexpressing the endogenous D-lactate dehydrogenase (D-LDH) detoxify D-lactate to pyruvate and survive |
| GO:0050660 flavin adenine dinucleotide binding | IBA GO_REF:0000033 | KEEP AS NON CORE | Summary: Flavin adenine dinucleotide binding: the enzyme carries a PCMH-type FAD-binding domain and uses FAD as the hydride acceptor; mouse LDHD crystal structures are FAD-bound and the reaction is defined with FAD as cosubstrate [PMID:37863926]. Reason: FAD binding is real and required (the catalytic reaction is written with FAD), but it is a cofactor-binding property subsumed by the FAD-dependent dehydrogenase activity term; kept as non-core so the core function remains the dehydrogenase activity. Supporting Evidence: file:DANRE/ldhd/ldhd-uniprot.txt FAD-binding PCMH-type file:DANRE/ldhd/ldhd-uniprot.txt Reaction=(R)-lactate + FAD + H(+) = FADH2 + pyruvate PMID:37863926 crystal structures of mLDHD in FAD-bound |
| GO:0140170 D-lactate dehydrogenase (FAD) activity | IEA GO_REF:0000120 | ACCEPT | Summary: D-lactate dehydrogenase (FAD) activity (Rhea:82479-mapped IEA): FAD-dependent oxidation of D-lactate to pyruvate. Zebrafish ldhd mutants accumulate D-lactate specifically and are rescued by human LDHD [PMID:30931947]; purified mouse and human LDHD oxidize D-lactate with FAD/Mn2+ and transfer electrons to cytochrome c [PMID:37863926; PMID:38413804]. Reason: This is the core molecular function. The zebrafish in vivo readout (D- but not L-lactate elevation, rescued by wild-type but not patient-variant human LDHD) demonstrates the enzyme's stereospecific activity in the fish, and the reaction chemistry (FAD as acceptor, Mn2+ cofactor) is established on the mammalian orthologs. Supporting Evidence: PMID:30931947 metabolic analysis revealed elevated levels of D-lactate, but not L-lactate, in ldhd−/− larvae compared to wildtype PMID:30931947 establishing that LDHD is stereospecific for D-lactate only file:DANRE/ldhd/ldhd-uniprot.txt Reaction=(R)-lactate + FAD + H(+) = FADH2 + pyruvate PMID:37863926 mLDHD is an Mn2+-dependent general dehydrogenase for a broad range of D-2-hydroxyacids with small to moderate-size hydrophobic moieties at the C2 atom |
| GO:0140174 (2R)-2-hydroxycarboxylate dehydrogenase (FAD) activity | IEA GO_REF:0000120 | ACCEPT | Summary: (2R)-2-hydroxycarboxylate dehydrogenase (FAD) activity (Rhea-mapped IEA (RHEA:82483-82511, 84927)): the broader substrate class. Mouse LDHD acts on D-2-hydroxybutanoate through D-2-hydroxyoctanoate and branched D-2-hydroxyacids with hydrophobic C2 groups, but not on hydrophilic ones or L-isomers [PMID:37863926]; human LDHD reduces cytochrome c with D-2-hydroxybutyrate as well [PMID:38413804]. Reason: D-lactate is one member of the substrate class this enzyme handles; the generic (2R)-2-hydroxycarboxylate term captures the true substrate scope shown biochemically on the mammalian orthologs, and the zebrafish enzyme is the ortholog whose D-lactate activity is confirmed in vivo. Accepted alongside the D-lactate-specific term. Supporting Evidence: file:DANRE/ldhd/ldhd-uniprot.txt Reaction=a (2R)-2-hydroxycarboxylate + FAD + H(+) = a 2-oxocarboxylate PMID:37863926 mLDHD is an Mn2+-dependent general dehydrogenase for a broad range of D-2-hydroxyacids with small to moderate-size hydrophobic moieties at the C2 atom PMID:38413804 we identified d-2-hydroxybutyrate as another LDHD substrate that can be used to reduce cytochrome c |
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Download this section (compressed HTML)Q: Does zebrafish Ldhd, like human LDHD, use cytochrome c as its physiological electron acceptor and contribute to mitochondrial membrane potential, or is its role in fish purely detoxification of D-lactate?
Experiment: Assay cytochrome c reduction by purified zebrafish Ldhd with D-lactate and D-2-hydroxybutyrate, and measure D-lactate-driven membrane potential in permeabilized cells from ldhd sa15623 versus wild-type larvae.
Hypothesis: Zebrafish Ldhd retains the cytochrome c-coupled electron transfer demonstrated for human LDHD.
Type: biochemical
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