IDH3B encodes the beta subunit of mitochondrial NAD-dependent isocitrate dehydrogenase 3 (IDH3). The functional IDH3 enzyme is a heterotetramer with alpha2-beta-gamma stoichiometry (2alpha:1beta:1gamma), where the alpha subunits (IDH3A) are catalytic, while beta (IDH3B) and gamma (IDH3G) play structural and regulatory roles. IDH3B does NOT have intrinsic catalytic activity on its own - expression of beta subunit alone shows no detectable activity [PMID:10601238]. The alphabeta and alphagamma dimers exhibit significant IDH activity, but the full heterotetramer is required for maximal activity and proper allosteric regulation [PMID:14555658]. The IDH3 complex catalyzes the irreversible oxidative decarboxylation of isocitrate to alpha-ketoglutarate in the TCA cycle. Beta-Arg99 is required for normal ADP activation of the enzyme but is not essential for catalysis [PMID:14555658]. IDH3B localizes to the mitochondrial matrix. Biallelic variants cause retinitis pigmentosa 46 (RP46). IDH3B exists as multiple isoforms (beta1 and beta2) due to alternative splicing, with tissue-specific expression patterns affecting the pH optimum of the enzyme complex [PMID:10601238].
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005739 mitochondrion | IBA GO_REF:0000033 | ACCEPT | Summary: IDH3B is well-established as a mitochondrial protein. The IBA annotation is phylogenetically well-supported across eukaryotes. UniProt indicates a mitochondrial transit peptide (residues 1-34) and documents mitochondrial localization. PMID:10601238 describes IDH3B as part of "mitochondrial NAD(+)-dependent isocitrate dehydrogenase" and discusses its function in the context of mitochondrial enzyme complexes. Reason: Mitochondrial localization is correct and well-supported by phylogenetic inference, UniProt annotation, and experimental literature. While mitochondrial matrix (GO:0005759) is more precise, this broader term is acceptable for the IBA evidence type. Supporting Evidence: PMID:10601238 the interactions and functional role of each of the three mitochondrial NAD(+)-dependent isocitrate dehydrogenase (IDH) subunits (alpha, beta, and gamma) |
| GO:0006099 tricarboxylic acid cycle | IBA GO_REF:0000033 | ACCEPT | Summary: IDH3B is a component of the NAD-dependent isocitrate dehydrogenase complex that catalyzes a key step of the TCA cycle. While the beta subunit is not itself catalytic, it is essential for proper complex assembly and function in the TCA cycle [PMID:10601238, PMID:14555658]. The IBA annotation appropriately captures the involvement of IDH3B in this process. Reason: As an essential structural component of the IDH3 complex, IDH3B participates in the TCA cycle even though it lacks intrinsic catalytic activity. The phylogenetic evidence supports this annotation across eukaryotes. Supporting Evidence: PMID:10601238 Among the various combinations of human IDH subunits co-expressed in bacteria, alphabetagamma, alphabeta, and alphagamma combinations exhibited significant amounts of IDH activity |
| GO:0006102 isocitrate metabolic process | IBA GO_REF:0000033 | ACCEPT | Summary: IDH3B participates in isocitrate metabolism as part of the IDH3 complex. The complex converts isocitrate to alpha-ketoglutarate. While IDH3B is not the catalytic subunit, it is required for proper complex assembly and activity [PMID:10601238]. Reason: The annotation correctly captures IDH3B's involvement in isocitrate metabolism through its role in the IDH3 complex. Well-supported by phylogenetic analysis. Supporting Evidence: PMID:10601238 the alpha is the catalytic subunit and that at least one of the other two subunits plays an essential supporting role for activity |
| GO:0000287 magnesium ion binding | IEA GO_REF:0000002 | MARK AS OVER ANNOTATED | Summary: This annotation is inferred from InterPro domain IPR019818 (IsoCit/isopropylmalate_DH_CS). However, in the IDH3 complex, Mg2+ (or Mn2+) binding occurs at the active site in the catalytic alpha subunit, not in the structural beta subunit. The deep research notes that the enzyme "is a heterooctamer containing two copies of a heterotetramer of two IDH3A, one IDH3B, one IDH3G, and two Mn++" [Reactome:R-HSA-70967], suggesting metal binding is associated with the catalytic subunits. No direct evidence supports magnesium binding by IDH3B specifically. The crystal structures of the αβ heterodimer (PDB 6KDE Ca2+-bound, 6KDY NAD-bound) place the divalent metal site at the active-site cleft contributed by the α subunit; the β subunit does not provide a productive metal site, and in the αβ heterodimer the active site has a distorted geometry unable to bind the metal ion in a catalysis-relevant manner [PMID:31515270]. Reason: While the InterPro domain is present, the functional magnesium/metal binding site is located in the catalytic alpha subunit. IDH3B shares sequence/structural homology with dehydrogenases but its specific role is structural, not catalytic. This IEA annotation represents domain-based inference that does not reflect the actual function of this specific subunit. Supporting Evidence: PMID:31515270 the active site has a distorted geometry that is unable to bind the metal ion effectively or in a catalysis-relevant manner |
| GO:0005739 mitochondrion | IEA GO_REF:0000044 | ACCEPT | Summary: Mitochondrial localization inferred from UniProt subcellular location annotation. This is consistent with experimental evidence and the IBA annotation above. Reason: Correct annotation based on UniProt curation. IDH3B has a mitochondrial transit peptide and functions in the mitochondrial matrix. |
| GO:0006099 tricarboxylic acid cycle | IEA GO_REF:0000120 | ACCEPT | Summary: Combined automated annotation correctly identifies IDH3B's involvement in the TCA cycle. Consistent with IBA annotation and experimental literature. Reason: Correct annotation. IDH3B is part of the IDH3 complex that catalyzes a key TCA cycle step. |
| GO:0016616 oxidoreductase activity, acting on the CH-OH group of donors, NAD or NADP as acceptor | IEA GO_REF:0000002 | REMOVE | Summary: This annotation is inferred from InterPro domain IPR019818. However, PMID:10601238 demonstrates that "subunits produced alone ... showed no detectable activity" and establishes that "the alpha is the catalytic subunit." IDH3B shares the dehydrogenase domain but does not itself possess oxidoreductase activity - it is a structural/ regulatory subunit of the complex. Reason: IDH3B does not have intrinsic oxidoreductase activity. The beta subunit alone shows no detectable enzymatic activity [PMID:10601238]. This annotation incorrectly attributes the catalytic activity of the complex to the non-catalytic beta subunit based solely on domain homology. Supporting Evidence: PMID:10601238 subunits produced alone and betagamma showed no detectable activity. These data suggest that the alpha is the catalytic subunit |
| GO:0051287 NAD binding | IEA GO_REF:0000002 | MARK AS OVER ANNOTATED | Summary: This annotation is inferred from InterPro domain IPR019818. While IDH3B shares structural homology with NAD-binding dehydrogenases, the functional NAD binding and catalysis occurs in the alpha subunit. PMID:14555658 shows that mutations in the gamma subunit affect NAD binding (gamma-R97Q has 10-fold higher Km for NAD), but the beta subunit's role is in ADP activation, not NAD binding at the catalytic site. However, the beta subunit may contribute to the nucleotide binding pocket architecture. Reason: The annotation is based on domain homology rather than demonstrated function. IDH3B contributes to the complex structure and ADP-mediated regulation but is not itself the primary NAD-binding subunit. The catalytic NAD binding site is in IDH3A. Crystal structures of the alphabeta heterodimer (PDB 6KDY NAD-bound, 6KE3 NADH-bound) show that the NAD cofactor binds in the active-site cleft built jointly from the alpha subunit and the small domain of the beta subunit, so the beta subunit contributes to the architecture of the holoenzyme NAD-binding/active site rather than binding NAD as an isolated subunit [PMID:31515270]. Supporting Evidence: PMID:14555658 the beta and gamma subunits have roles in the nucleotide functions of this allosteric enzyme PMID:31515270 We report here the crystal structures of the αβ heterodimer of human NAD-IDH with the α subunit in apo form and in Ca2+-bound, NAD-bound, and NADH-bound forms PMID:31515270 The active site is located in the cleft formed by the large and small domains of the α subunit and the small domain of the β subunit PMID:36375638 the αQ139AICT+Ca+NADβNAD structure presents the first pseudo-Michaelis complex of HsIDH3, which allows us to identify the key residues involved in the binding of cofactor, substrate, and metal ion |
| GO:0045242 isocitrate dehydrogenase complex (NAD+) | IEA GO_REF:0000107 | ACCEPT | Summary: IDH3B is a core component of the NAD-dependent isocitrate dehydrogenase complex. The complex has 2alpha:1beta:1gamma stoichiometry [PMID:14555658]. This cellular component annotation is entirely appropriate for IDH3B. Crystal structures of the human IDH3 alphabeta heterodimer (PDB 6KDE/6KDF/6KDY/6KE3) and of the constitutively active (αβαγ)2 holoenzyme (PDB 8GRB) directly establish IDH3B (beta) as an assembled subunit of the IDH3 complex and define the alpha-beta heterodimer interface [PMID:31515270, PMID:36375638]. Reason: Correct annotation. IDH3B is an established subunit of the IDH3 complex, confirmed by co-expression studies and structural analysis. Supporting Evidence: PMID:14555658 Wild type (WT) and mutant enzymes ... were purified to homogeneity yielding enzymes with 2alpha:1beta:1gamma subunit composition PMID:31515270 The α and β subunits form a heterodimer (αβ), and the α and γ subunits form another heterodimer (αγ), which are assembled into a heterotetramer (α2βγ) and further into a heterooctamer PMID:36375638 It consists of three types of subunits (α, β, and γ) and exists and functions as the (αβαγ)2 heterooctamer |
| GO:0005739 mitochondrion | IDA GO_REF:0000052 | ACCEPT | Summary: Direct experimental evidence for mitochondrial localization from HPA immunofluorescence data. This provides direct experimental support for the mitochondrial localization. Reason: Direct experimental evidence (IDA from immunofluorescence) confirms mitochondrial localization, consistent with all other evidence sources. |
| GO:0005739 mitochondrion | NAS PMID:14555658 Evaluation by mutagenesis of the importance of 3 arginines i... | ACCEPT | Summary: PMID:14555658 is a study of human NAD-dependent isocitrate dehydrogenase expressed in bacteria. The paper discusses the enzyme's function in the context of mitochondrial metabolism but does not directly demonstrate mitochondrial localization - hence NAS (non-traceable author statement) is appropriate. Reason: While the evidence is indirect, mitochondrial localization is well-established from multiple other sources. This annotation is consistent with the overall body of evidence. Supporting Evidence: PMID:14555658 Mammalian NAD-dependent isocitrate dehydrogenase is an allosteric enzyme |
| GO:0006099 tricarboxylic acid cycle | IDA PMID:14555658 Evaluation by mutagenesis of the importance of 3 arginines i... | ACCEPT | Summary: PMID:14555658 demonstrates that the IDH3 complex including the beta subunit has isocitrate dehydrogenase activity, which is a key TCA cycle reaction. The study shows IDH activity of purified complexes containing beta subunit [PMID:14555658]. Reason: Direct experimental evidence that IDH3B-containing complexes have isocitrate dehydrogenase activity, supporting involvement in TCA cycle. Supporting Evidence: PMID:14555658 Specific activities of 22, 14, and 2 micromol of NADH/min/mg were measured, respectively, for WT, beta-R99Q, and gamma-R97Q enzymes |
| GO:0005739 mitochondrion | HTP PMID:34800366 Quantitative high-confidence human mitochondrial proteome an... | ACCEPT | Summary: PMID:34800366 is a high-throughput mitochondrial proteomics study that identified IDH3B in the human mitochondrial proteome. This provides orthogonal mass spectrometry- based evidence for mitochondrial localization. Reason: High-quality proteomics study confirming IDH3B presence in the mitochondrial proteome. Supporting Evidence: PMID:34800366 Quantitative high-confidence human mitochondrial proteome |
| GO:0045242 isocitrate dehydrogenase complex (NAD+) | IDA PMID:14555658 Evaluation by mutagenesis of the importance of 3 arginines i... | ACCEPT | Summary: PMID:14555658 provides direct experimental evidence that IDH3B is a component of the IDH3 complex. The study purified the complex and demonstrated the 2alpha:1beta:1gamma stoichiometry through expression and biochemical analysis. Crystal structures of the human IDH3 alphabeta heterodimer (PDB 6KDE/6KDF/6KDY NAD-bound/6KE3 NADH-bound) and of the constitutively active (alphabeta-alphagamma)2 holoenzyme (PDB 8GRB) directly visualize IDH3B (beta) as an assembled subunit of the IDH3 complex and define the alpha-beta heterodimer interface [PMID:31515270, PMID:36375638]. Reason: Direct experimental demonstration that IDH3B is part of the IDH3 complex with defined stoichiometry. Supporting Evidence: PMID:14555658 Wild type (WT) and mutant enzymes (each containing 2 normal subunits plus a mutant subunit with alpha-R88Q, beta-R99Q, or gamma-R97Q) were purified to homogeneity yielding enzymes with 2alpha:1beta:1gamma subunit composition and a native molecular mass of 315 kDa PMID:31515270 The α and β subunits form a heterodimer (αβ), and the α and γ subunits form another heterodimer (αγ), which are assembled into a heterotetramer (α2βγ) and further into a heterooctamer PMID:36375638 It consists of three types of subunits (α, β, and γ) and exists and functions as the (αβαγ)2 heterooctamer |
| GO:0005634 nucleus | HDA PMID:21630459 Proteomic characterization of the human sperm nucleus. | KEEP AS NON CORE | Summary: PMID:21630459 is a proteomic study of the human sperm nucleus that identified IDH3B among 403 nuclear proteins. This is a specialized cell type where metabolic proteins may be present in the nuclear fraction during spermatogenesis. The nuclear localization is likely context-specific rather than representing a core localization. Reason: The detection in sperm nuclear proteome may represent a specialized developmental context. IDH3B's primary localization is mitochondrial, and nuclear detection may reflect the unique chromatin remodeling during spermiogenesis or technical co-purification. This is not a core localization but should be retained. Supporting Evidence: PMID:21630459 403 different proteins have been identified from the isolated sperm nuclei |
| GO:0005759 mitochondrial matrix | TAS Reactome:R-HSA-70967 | ACCEPT | Summary: Reactome pathway R-HSA-70967 "IDH3 complex decarboxylates isocitrate" places the IDH3 complex, including IDH3B, in the mitochondrial matrix. This is the most specific and accurate localization for IDH3B's function. Reason: Mitochondrial matrix is the precise subcellular location where IDH3 functions in the TCA cycle. This is the most informative localization annotation for IDH3B. |
| GO:0009055 electron transfer activity | TAS PMID:10601238 Identification and functional characterization of a novel, t... | REMOVE | Summary: PMID:10601238 does not support electron transfer activity for IDH3B. The paper demonstrates that the IDH3 complex has isocitrate dehydrogenase activity (reducing NAD+ to NADH), but this is not the same as electron transfer activity. Electron transfer activity (GO:0009055) refers to proteins that function as electron carriers (like cytochromes or ferredoxins), not dehydrogenases that use NAD+ as a cofactor. Furthermore, IDH3B specifically is the non-catalytic subunit. Reason: Incorrect annotation. GO:0009055 (electron transfer activity) is inappropriate for IDH3B. The NAD+-dependent dehydrogenase reaction is not electron transfer activity in the GO sense. Additionally, IDH3B is not catalytically active - the paper states "subunits produced alone ... showed no detectable activity." Supporting Evidence: PMID:10601238 subunits produced alone and betagamma showed no detectable activity |
| GO:0004449 isocitrate dehydrogenase (NAD+) activity | TAS PMID:10601238 Identification and functional characterization of a novel, t... | REMOVE | Summary: PMID:10601238 explicitly demonstrates that IDH3B alone does not have isocitrate dehydrogenase activity. The paper states "subunits produced alone and betagamma showed no detectable activity" and concludes that "the alpha is the catalytic subunit." While the alphabeta dimer has activity, this is due to the alpha subunit, not beta. Reason: The cited paper PMID:10601238 directly contradicts this annotation. IDH3B does not have intrinsic isocitrate dehydrogenase activity. The beta subunit is structural/ regulatory, and attributing the catalytic activity to it is incorrect. This is a case where the annotation was likely made at the complex level and incorrectly propagated to the non-catalytic subunit. Supporting Evidence: PMID:10601238 subunits produced alone and betagamma showed no detectable activity. These data suggest that the alpha is the catalytic subunit and that at least one of the other two subunits plays an essential supporting role for activity |
| GO:0005739 mitochondrion | TAS PMID:10601238 Identification and functional characterization of a novel, t... | ACCEPT | Summary: PMID:10601238 describes IDH3B as part of "mitochondrial NAD(+)-dependent isocitrate dehydrogenase" and discusses it in the context of mitochondrial function. The mitochondrial localization is well-supported. Reason: The paper describes IDH3B as a mitochondrial protein. Consistent with all other evidence for mitochondrial localization. Supporting Evidence: PMID:10601238 the interactions and functional role of each of the three mitochondrial NAD(+)-dependent isocitrate dehydrogenase (IDH) subunits |
| GO:0006102 isocitrate metabolic process | TAS PMID:10601238 Identification and functional characterization of a novel, t... | ACCEPT | Summary: PMID:10601238 demonstrates that IDH3B contributes to isocitrate metabolism as part of the IDH3 complex. While IDH3B is not catalytic, it plays "an essential supporting role for activity" in isocitrate oxidation. Reason: IDH3B participates in isocitrate metabolism through its essential structural role in the IDH3 complex. The paper demonstrates that beta-containing complexes have isocitrate dehydrogenase activity. Supporting Evidence: PMID:10601238 alphabetagamma, alphabeta, and alphagamma combinations exhibited significant amounts of IDH activity |
| GO:0005198 structural molecule activity | NAS PMID:10601238 Identification and functional characterization of a novel, t... | NEW | Summary: IDH3B functions as a structural subunit of the IDH3 complex. PMID:10601238 demonstrates that the beta subunit alone has no catalytic activity ("subunits produced alone and betagamma showed no detectable activity") but plays "an essential supporting role for activity." The beta subunit is required for proper complex assembly and function. Deep research confirms that IDH3B plays a structural/regulatory role in the heterotetramer [file:human/IDH3B/IDH3B-deep-research-falcon.md]. Reason: This annotation captures IDH3B's established role as a structural component of the IDH3 enzyme complex. The term GO:0005198 is defined as "The action of a molecule that contributes to the structural integrity of a complex" which accurately describes IDH3B's function. This is a more appropriate molecular function annotation than the catalytic activity terms that were incorrectly attributed to IDH3B. Supporting Evidence: PMID:10601238 These data suggest that the alpha is the catalytic subunit and that at least one of the other two subunits plays an essential supporting role for activity file:human/IDH3B/IDH3B-deep-research-falcon.md IDH3B appears to play a structural/regulatory role in the complex, as opposed to a catalytic one |
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Download this section (compressed HTML)Q: Within the αβαγ heterotetramer (and its (αβαγ)2 holoenzyme), which IDH3B residues beyond β-Arg99 transmit the ADP allosteric signal to the IDH3A catalytic site, and does IDH3B contribute any direct interactions to substrate/cofactor binding in the pseudo-Michaelis complex captured by the αQ139A·ICT·Ca·NAD·βNAD structure (PMID:36375638)? Mapping these residues would justify more specific molecular-function annotations for IDH3B (e.g., positive regulation of isocitrate dehydrogenase activity).
Q: What is the functional consequence of β1 vs β2 isoform switching in vivo? Beyond the shifted pH optimum from 8.0 to 7.6 in vitro (PMID:10601238), do heart and skeletal muscle preferentially run IDH3 at distinct intramitochondrial pH set points, and is this isoform exchange dynamic with physiological state (exercise, ischemia)?
Q: Why does loss of IDH3B selectively manifest as retinitis pigmentosa 46 when IDH3 operates ubiquitously? Is the retinal phenotype driven by a uniquely high photoreceptor TCA flux, by altered NADH/NAD+ balance affecting outer-segment lipid/visual-cycle pathways, or by α2γ-only complexes acting as a partial-loss-of- function background that other tissues tolerate?
Experiment: Reconstitute the recombinant (αβαγ)2 holoenzyme with IDH3B variants spanning the β3-α3 and β12-α8 loops identified in PMID:31515270 / PMID:36375638 as building the pseudo-allosteric site, and measure ADP-dependent k_cat / K_m for isocitrate plus ADP-binding affinity by ITC. Pair with cryo-EM of selected variants in the apo and ADP-bound states to confirm structural consequences.
Hypothesis: The pseudo-allosteric loops contributed by IDH3B (small domain plus β3-α3 / β12-α8) are the principal transducers of ADP activation, and engineered residue swaps in these loops can uncouple ADP binding from catalytic activation.
Type: biochemistry / structural biology
Experiment: Generate isoform-specific β1 and β2 CRISPR knock-in HEK293, C2C12 (skeletal muscle) and HL-1 (cardiomyocyte) lines and measure intramitochondrial pH, TCA flux (13C isotopologue analysis of citrate/α-ketoglutarate), and oxygen consumption under basal and exercised/hypoxic conditions.
Hypothesis: β2 expression in muscle tissue tunes IDH3 activity to a lower, physiologically relevant matrix pH, supporting sustained TCA flux during high-workload conditions where matrix pH falls below the β1 optimum.
Type: cell biology / metabolomics
Experiment: Patient-derived iPSC differentiation to retinal organoids from RP46 IDH3B patients vs. isogenic CRISPR-corrected controls, with assessment of photoreceptor survival, outer-segment morphology, NADH/NAD+ ratio, α-KG levels, and TCA flux. Compare to iPSC-derived cardiomyocytes and hepatocytes from the same patients to identify the metabolic axis that distinguishes the retina.
Hypothesis: Photoreceptor degeneration in RP46 is driven by NADH-dependent or α-KG-dependent processes uniquely sensitive to partial IDH3 activity (e.g., visual-cycle retinoid metabolism or HIF/2-oxoglutarate-dependent dioxygenase signalling) rather than by a generalized TCA failure.
Type: stem-cell biology / clinical model
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