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Garry K. Brown - One of the best experts on this subject based on the ideXlab platform.
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pyruvate dehydrogenase E3 Binding Protein Protein x deficiency
Developmental Medicine & Child Neurology, 2006Co-Authors: Ruth M. Brown, Rosie Head, A A M Morris, J A J Raiman, J H Walter, William P Whitehouse, Garry K. BrownAbstract:Pyruvate dehydrogenase (PDH) deficiency is a major cause of neurological dysfunction and lactic acidosis in infancy and early childhood. The great majority of cases (>80%) result from mutations in the X-linked gene for the E1α subunit of the complex (PDHA1). Mutations in the genes for the other subunits have all been described, but only dihydrolipoamide dehydrogenase (E3) and E3 Binding Protein (E3BP) defects contribute significantly to the total number of patients with PDH deficiency. Although previously considered rare, with only 13 reported cases, we have found that mutations in PDX1, the gene for the E3 Binding Protein, are in fact relatively common. Clinical, biochemical, and genetic features of six new patients (four males, two females; age range 15mo-6y) with mutations in this gene are compared with previously reported cases. All patients with E3BP deficiency identified to date have mutations which completely prevent synthesis of the Protein product. However, they are generally less severely affected than patients with PDHA1 mutations, although there is considerable overlap in clinical and neuroradiological features.
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Lactic acidosis and developmental delay due to deficiency of E3 Binding Protein (Protein X) of the pyruvate dehydrogenase complex
Journal of Inherited Metabolic Disease, 2004Co-Authors: D. G. Ramadan, Ruth M. Brown, Rosie A. Head, A. Al-tawari, Y. Habeeb, M. Zaki, F. Al-ruqum, G. T. N. Besley, J. E. Wraith, Garry K. BrownAbstract:: Pyruvate dehydrogenase deficiency is an important cause of primary lactic acidosis. Most cases occur as a result of mutations in the gene for the E1α subunit of the complex, with a small number resulting from mutations in genes for other components, most commonly the E3 and E3-Binding Protein subunits. We describe pyruvate dehydrogenase E3-Binding Protein deficiency in two siblings in each of two unrelated families from Kuwait. The index patient in each family had reduced pyruvate dehydrogenase activity in cultured fibroblasts and no detectable immunoreactive E3-Binding Protein. Both were homozygous for nonsense mutations in the E3-Binding Protein gene, one involving the codon for glutamine 266, the other the codon for tryptophan 5.
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Pyruvate dehydrogenase E3 Binding Protein deficiency
Human Genetics, 2002Co-Authors: Ruth M. Brown, Rosie A. Head, Garry K. BrownAbstract:Primary defects of the E3 Binding Protein component of the pyruvate dehydrogenase complex appear to be a rare cause of pyruvate dehydrogenase deficiency. We describe two new, unrelated patients with mutations in the E3 Binding Protein gene, in both cases involving the conserved dinucleotides of splice junctions. Both patients presented with delayed development and lactic acidosis, features that are also found in patients with the more common pyruvate dehydrogenase E1α subunit deficiency; however, they both had significant residual enzyme activity in cultured fibroblasts and prolonged survival.
Ruth M. Brown - One of the best experts on this subject based on the ideXlab platform.
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pyruvate dehydrogenase E3 Binding Protein Protein x deficiency
Developmental Medicine & Child Neurology, 2006Co-Authors: Ruth M. Brown, Rosie Head, A A M Morris, J A J Raiman, J H Walter, William P Whitehouse, Garry K. BrownAbstract:Pyruvate dehydrogenase (PDH) deficiency is a major cause of neurological dysfunction and lactic acidosis in infancy and early childhood. The great majority of cases (>80%) result from mutations in the X-linked gene for the E1α subunit of the complex (PDHA1). Mutations in the genes for the other subunits have all been described, but only dihydrolipoamide dehydrogenase (E3) and E3 Binding Protein (E3BP) defects contribute significantly to the total number of patients with PDH deficiency. Although previously considered rare, with only 13 reported cases, we have found that mutations in PDX1, the gene for the E3 Binding Protein, are in fact relatively common. Clinical, biochemical, and genetic features of six new patients (four males, two females; age range 15mo-6y) with mutations in this gene are compared with previously reported cases. All patients with E3BP deficiency identified to date have mutations which completely prevent synthesis of the Protein product. However, they are generally less severely affected than patients with PDHA1 mutations, although there is considerable overlap in clinical and neuroradiological features.
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Lactic acidosis and developmental delay due to deficiency of E3 Binding Protein (Protein X) of the pyruvate dehydrogenase complex
Journal of Inherited Metabolic Disease, 2004Co-Authors: D. G. Ramadan, Ruth M. Brown, Rosie A. Head, A. Al-tawari, Y. Habeeb, M. Zaki, F. Al-ruqum, G. T. N. Besley, J. E. Wraith, Garry K. BrownAbstract:: Pyruvate dehydrogenase deficiency is an important cause of primary lactic acidosis. Most cases occur as a result of mutations in the gene for the E1α subunit of the complex, with a small number resulting from mutations in genes for other components, most commonly the E3 and E3-Binding Protein subunits. We describe pyruvate dehydrogenase E3-Binding Protein deficiency in two siblings in each of two unrelated families from Kuwait. The index patient in each family had reduced pyruvate dehydrogenase activity in cultured fibroblasts and no detectable immunoreactive E3-Binding Protein. Both were homozygous for nonsense mutations in the E3-Binding Protein gene, one involving the codon for glutamine 266, the other the codon for tryptophan 5.
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Pyruvate dehydrogenase E3 Binding Protein deficiency
Human Genetics, 2002Co-Authors: Ruth M. Brown, Rosie A. Head, Garry K. BrownAbstract:Primary defects of the E3 Binding Protein component of the pyruvate dehydrogenase complex appear to be a rare cause of pyruvate dehydrogenase deficiency. We describe two new, unrelated patients with mutations in the E3 Binding Protein gene, in both cases involving the conserved dinucleotides of splice junctions. Both patients presented with delayed development and lactic acidosis, features that are also found in patients with the more common pyruvate dehydrogenase E1α subunit deficiency; however, they both had significant residual enzyme activity in cultured fibroblasts and prolonged survival.
Thomas E Roche - One of the best experts on this subject based on the ideXlab platform.
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organization of the cores of the mammalian pyruvate dehydrogenase complex formed by e2 and e2 plus the E3 Binding Protein and their capacities to bind the e1 and E3 components
Journal of Biological Chemistry, 2004Co-Authors: Yasuaki Hiromasa, Tetsuro Fujisawa, Thomas E RocheAbstract:Abstract The subunits of the dihydrolipoyl acetyltransferase (E2) component of mammalian pyruvate dehydrogenase complex can form a 60-mer via association of the C-terminal I domain of E2 at the vertices of a dodecahedron. Exterior to this inner core structure, E2 has a pyruvate dehydrogenase component (E1)-Binding domain followed by two lipoyl domains, all connected by mobile linker regions. The assembled core structure of mammalian pyruvate dehydrogenase complex also includes the dihydrolipoyl dehydrogenase (E3)-Binding Protein (E3BP) that binds the I domain of E2 by its C-terminal I′ domain. E3BP similarly has linker regions connecting an E3-Binding domain and a lipoyl domain. The composition of E2·E3BP was thought to be 60 E2 plus ∼12 E3BP. We have prepared homogenous human components. E2 and E2·E3BP have s20,w values of 36 S and 31.8 S, respectively. Equilibrium sedimentation and small angle x-ray scattering studies indicate that E2·E3BP has lower total mass than E2, and small angle x-ray scattering showed that E3 binds to E2·E3BP outside the central dodecahedron. In the presence of saturating levels of E1, E2 bound ∼60 E1 and maximally sedimented 64.4 ± 1.5 S faster than E2, whereas E1-saturated E2·E3BP maximally sedimented 49.5 ± 1.4 S faster than E2·E3BP. Based on the impact on sedimentation rates by bound E1, we estimate fewer E1 (∼12) were bound by E2·E3BP than by E2. The findings of a smaller E2·E3BP mass and a lower capacity to bind E1 support the smaller E3BP substituting for E2 subunits rather than adding to the 60-mer. We describe a substitution model in which 12 I′ domains of E3BP replace 12 I domains of E2 by forming 6 dimer edges that are symmetrically located in the dodecahedron structure. Twelve E3 dimers were bound per E248·E3BP12 mass, which is consistent with this model.
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Facilitated Interaction between the Pyruvate Dehydrogenase Kinase Isoform 2 and the Dihydrolipoyl Acetyltransferase
Journal of Biological Chemistry, 2003Co-Authors: Yasuaki Hiromasa, Thomas E RocheAbstract:Abstract The dihydrolipoyl acetyltransferase (E2) has an enormous impact on pyruvate dehydrogenase kinase (PDK) phosphorylation of the pyruvate dehydrogenase (E1) component by acting as a mobile Binding framework and in facilitating and mediating regulation of PDK activity. Analytical ultracentrifugation (AUC) studies established that the soluble PDK2 isoform is a stable dimer. The interaction of PDK2 with the lipoyl domains of E2 (L1, L2) and the E3-Binding Protein (L3) were characterized by AUC. PDK2 interacted very weakly with L2 (Kd ≃ 175 μm for 2 L2/PDK2) but much tighter with dimeric glutathione S-transferase (GST)-L2 (Kd ≃ 3 μm), supporting the importance of bifunctional Binding. Reduction of lipoyl groups resulted in ∼8-fold tighter Binding of PDK2 to GST-L2red, which was ∼300-fold tighter than Binding of 2 L2red and also much tighter than Binding by GST-L1red and GST-L3red. The E2 60-mer bound ∼18 PDK2 dimers with a Kd similar to GST-L2. E2·E1 bound more PDK2 (∼27.6) than E2 with ∼2-fold tighter affinity. Lipoate reduction fostered somewhat tighter Binding at more sites by E2 and severalfold tighter Binding at the majority of sites on E2·E1. ATP and ADP decreased the affinity of PDK2 for E2 by 3–5-fold and adenosine 5′-(β,γ-imino)triphosphate or phosphorylation of E1 similarly reduced PDK2 Binding to E2·E1. Reversible bifunctional Binding to L2 with the mandatory singly held transition fits the proposed “hand-over-hand” movement of a kinase dimer to access E1 without dissociating from the complex. The gain in Binding interactions upon lipoate reduction likely aids reduction-engendered stimulation of PDK2 activity; loosening of Binding as a result of adenine nucleotides and phosphorylation may instigate movement of lipoyl domain-held kinase to a new E1 substrate.
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Assembly and Full Functionality of Recombinantly Expressed Dihydrolipoyl Acetyltransferase Component of the Human Pyruvate Dehydrogenase Complex
Journal of Biological Chemistry, 1997Co-Authors: Daqing Yang, Jiasheng Song, T. Wagenknecht, Thomas E RocheAbstract:Abstract The dihydrolipoyl acetyltransferase (E2) component of mammalian pyruvate dehydrogenase complex (PDC) consists of 60 COOH-terminal domains as an inner assemblage and sequentially via linker regions an exterior pyruvate dehydrogenase (E1) Binding domain and two lipoyl domains. Mature human E2, expressed in a protease-deficient Escherichia coli strain at 27°, was prepared in a highly purified form. Purified E2 had a high acetyltransferase activity, was well lipoylated based on its acetylation, and bound a large complement of bovine E1. Electron micrographs demonstrated that the inner core was assembled in the expected pentagonal dodecahedron shape with E1 Binding around the inner core periphery. With saturating E1 and excess dihydrolipoyl dehydrogenase (E3) but no E3-Binding Protein (E3BP), the recombinant E2 supported the overall PDC reaction at 4% of the rate of bovine E2·E3BP subcomplex. The lipoates of assembled human E2 or its free bilipoyl domain region were reduced by E3 at rates proportional to the lipoyl domain concentration, but those of the E2·E3BP were rapidly used in a concentration-independent manner consistent with bound E3 rapidly using a set of lipoyl domains localized nearby. Given this restriction and the need for E3BP for high PDC activity, directed channeling of reducing equivalents to bound E3 must be very efficient in the complex. The recombinant E2 oligomer increased E1 kinase activity by up to 4-fold and, in a Ca2+-dependent process, increased phospho-E1 phosphatase activity more than 15-fold. Thus the E2 assemblage fully provides the molecular intervention whereby a single E2-bound kinase or phosphatase molecule rapidly phosphorylate or dephosphorylate, respectively, many E2-bound E1. Thus, we prepared properly assembled, fully functional human E2 that mediated enhanced regulatory enzyme activities but, lacking E3BP, supported low PDC activity.
Peter W. Stacpoole - One of the best experts on this subject based on the ideXlab platform.
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The spectrum of pyruvate dehydrogenase complex deficiency: clinical, biochemical and genetic features in 371 patients.
Molecular genetics and metabolism, 2020Co-Authors: Kavi P. Patel, Jonathan Shuster, Sankarasubramon H. Subramony, Thomas W. O'brien, Peter W. StacpooleAbstract:Pyruvate dehydrogenase complex (PDC) deficiency is a genetic mitochondrial disorder commonly associated with lactic acidosis, progressive neurological and neuromuscular degeneration and, usually, death during childhood. There has been no recent comprehensive analysis of the natural history and clinical course of this disease. We reviewed 371 cases of PDC deficiency, published between 1970 and 2010, that involved defects in subunits E1α and E1β and components E1, E2, E3 and the E3 Binding Protein of the complex. English language peer-reviewed publications were identified, primarily by using PubMed and Google Scholar search engines. Neurodevelopmental delay and hypotonia were the commonest clinical signs of PDC deficiency. Structural brain abnormalities frequently included ventriculomegaly, dysgenesis of the corpus callosum and neuroimaging findings typical of Leigh syndrome. Neither gender nor any clinical or neuroimaging feature differentiated the various biochemical etiologies of the disease. Patients who died were younger, presented clinically earlier and had higher blood lactate levels and lower residual enzyme activities than subjects who were still alive at the time of reporting. Survival bore no relationship to the underlying biochemical or genetic abnormality or to gender. Although the clinical spectrum of PDC deficiency is broad, the dominant clinical phenotype includes presentation during the first year of life; neurological and neuromuscular degeneration; structural lesions revealed by neuroimaging; lactic acidosis and a blood lactate:pyruvate ratio ≤ 20.
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The spectrum of pyruvate dehydrogenase complex deficiency: Clinical, biochemical and genetic features in 371 patients
Molecular Genetics and Metabolism, 2012Co-Authors: Kavi P. Patel, Jonathan Shuster, Sankarasubramon H. Subramony, Thomas W. O'brien, Peter W. StacpooleAbstract:Context: Pyruvate dehydrogenase complex (PDC) deficiency is a genetic mitochondrial disorder commonly associated with lactic acidosis, progressive neurological and neuromuscular degeneration and, usually, death during childhood. There has been no recent comprehensive analysis of the natural history and clinical course of this disease. Objective: We reviewed 371 cases of PDC deficiency, published between 1970 and 2010, that involved defects in subunits E1α and E1β and components E1, E2, E3 and the E3 Binding Protein of the complex. Data sources and extraction: English language peer-reviewed publications were identified, primarily by using PubMed and Google Scholar search engines. Results: Neurodevelopmental delay and hypotonia were the commonest clinical signs of PDC deficiency. Structural brain abnormalities frequently included ventriculomegaly, dysgenesis of the corpus callosum and neuroimaging findings typical of Leigh syndrome. Neither gender nor any clinical or neuroimaging feature differentiated the various biochemical etiologies of the disease. Patients who died were younger, presented clinically earlier and had higher blood lactate levels and lower residual enzyme activities than subjects who were still alive at the time of reporting. Survival bore no relationship to the underlying biochemical or genetic abnormality or to gender. Conclusions: Although the clinical spectrum of PDC deficiency is broad, the dominant clinical phenotype includes presentation during the first year of life; neurological and neuromuscular degeneration; structural lesions revealed by neuroimaging; lactic acidosis and a blood lactate:pyruvate ratio ≤ 20. © 2012 Elsevier Inc.
Stephen J. Yeaman - One of the best experts on this subject based on the ideXlab platform.
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Autoreactive responses to pyruvate dehydrogenase complex in the pathogenesis of primary biliary cirrhosis
Immunological Reviews, 2000Co-Authors: Stephen J. Yeaman, John A. Kirby, David JonesAbstract:Primary biliary cirrhosis (PBC) is a cholestatic liver disease characterised by immune-mediated destruction of the biliary epithelial cells (BEC) lining the intrahepatic bile ducts (non-suppurative destructive cholangitis (NSDC)). Autoantibody and autoreactive T-cell responses specific for the self-antigen pyruvate dehydrogenase complex (PDC) are almost ubiquitous in PBC patients, leading to the view that the disease has an autoimmune aetiology. Autoreactive responses in PBC appear to be directed at the E2 and at the E3-Binding Protein (E3BP) (Protein X) components of PDC, with the dominant B-cell and T-cell epitopes in E2 (fewer data are available for E3BP) spanning the inner (of two) lipoic acid-Binding domains. The causal link between the breakdown of self-tolerance to PDC (particularly at the T-cell level) and the development of NSDC has been emphasised by the demonstration, in a murine model (experimental autoimmune cholangitis), that sensitisation with PDC of mammalian origin results in a breakdown of both B-cell and T-cell tolerance to murine PDC accompanied by the development of NSDC. An increasing understanding of the role played by PDC-specific autoreactive T cells in the pathogenesis of PBC has led us to examine the role played by the target cells in PBC (BEC) in both the inducer and effector mechanisms responsible for PBC.
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Characterization of the autoantibody responses to recombinant E3 Binding Protein (Protein X) of pyruvate dehydrogenase in primary biliary cirrhosis
Hepatology, 1999Co-Authors: Jeremy M. Palmer, David Jones, Janet Quinn, Anna Mchugh, Stephen J. YeamanAbstract:Autoantibodies to the pyruvate dehydrogenase complex (PDC) are present in the serum of more than 95% of patients with primary biliary cirrhosis (PBC), the major epitope being the inner lipoyl domain of the E2 component. Immunoblotting suggests a similar prevalence of antibodies to a tightly associated lipoic acid–containing Protein, E3 Binding Protein (;E3BP). Attempts to resolve E3BP from E2 have been unsuccessful, restricting study of the nature and significance of antibody responses to the individual Proteins. In particular, it is unclear (;1) whether there is true cross-reactivity between E3BP and E2 and, if so, which is the originating response and (;2) whether autoantibodies preferentially bind a lipoylated epitope on E3BP as is the case with PDC-E2. In this study, complementary DNAs encoding rE2, full-length rE3BP, its single lipoyl domain (;rLip), and core domain (;rE3BPCore) were cloned, and the Proteins were expressed in Escherichia coli. Sera from 47 PBC patients were studied by immunoblotting and enzyme-linked immunosorbent assay (;ELISA) against rE2, rE3BP, rE3BPCore, and both unlipoylated (;U) and lipoylated (;L) rLip. All sera were reactive by ELISA to some degree with all recombinant Proteins except rE3BPCore, to which only 6 of 47 showed any reactivity. Significant correlations (;P
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characterization of the autoantibody responses to recombinant E3 Binding Protein Protein x of pyruvate dehydrogenase in primary biliary cirrhosis
Hepatology, 1999Co-Authors: Jeremy M. Palmer, David Jones, Janet Quinn, Anna Mchugh, Stephen J. YeamanAbstract:Autoantibodies to the pyruvate dehydrogenase complex (PDC) are present in the serum of more than 95% of patients with primary biliary cirrhosis (PBC), the major epitope being the inner lipoyl domain of the E2 component. Immunoblotting suggests a similar prevalence of antibodies to a tightly associated lipoic acid–containing Protein, E3 Binding Protein (;E3BP). Attempts to resolve E3BP from E2 have been unsuccessful, restricting study of the nature and significance of antibody responses to the individual Proteins. In particular, it is unclear (;1) whether there is true cross-reactivity between E3BP and E2 and, if so, which is the originating response and (;2) whether autoantibodies preferentially bind a lipoylated epitope on E3BP as is the case with PDC-E2. In this study, complementary DNAs encoding rE2, full-length rE3BP, its single lipoyl domain (;rLip), and core domain (;rE3BPCore) were cloned, and the Proteins were expressed in Escherichia coli. Sera from 47 PBC patients were studied by immunoblotting and enzyme-linked immunosorbent assay (;ELISA) against rE2, rE3BP, rE3BPCore, and both unlipoylated (;U) and lipoylated (;L) rLip. All sera were reactive by ELISA to some degree with all recombinant Proteins except rE3BPCore, to which only 6 of 47 showed any reactivity. Significant correlations (;P < .0001) were observed when comparing absorbance values for rE3BP with both rLip (;U) (;r = 0.793) and (;L) (;r = 0.963). The mean absorbance for rLip (;U, 0.26 ± 0.05) was, however, significantly lower than the absorbance for rLip (;L) (;0.78 ± 0.12; P < .0001). After probing by immunoblotting and elution of antibodies from rE2 and rE3BP, subsequent reprobing against the components in whole PDC revealed true cross-reactivity. In summary, the response to E3BP is primarily directed against the lipoylated domain of the Protein. It still remains unclear, however, whether the initial breakdown of tolerance is to E2 or E3BP.
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T cell responses to the putative dominant autoepitope in primary biliary cirrhosis (PBC)
Clinical and Experimental Immunology, 1999Co-Authors: Jeremy M. Palmer, Stephen J. Yeaman, A. G. Diamond, Margaret F. Bassendine, David JonesAbstract:PBC is characterized by T cell-mediated destruction of the biliary epithelial cells lining the small intrahepatic bile ducts. The E2 and E3 Binding Protein (E3BP (Protein X)) components of pyruvate dehydrogenase complex (PDC) are disease-specific autoantigens in PBC. Attempts to localize the T cell autoepitopes within PDC-E2 have, however, generated contradictory results. One study has suggested the presence of T cell epitopes throughout PDC-E2, whilst another has identified a single dominant 14 amino acid T cell epitope (p163) spanning the lipoic acid Binding lysine residue in the inner lipoyl domain (ILD) of PDC-E2. The aim of the current study was to determine the prevalence of T cell responses to p163 and PDC-E2 ILD, and the role played by lipoylation of these antigens in their immunogenicity, in a UK PBC population. We found that the majority of the PBC patients showing a 6-day peripheral blood T cell proliferative response to native human PDC also responded, in a MHC class II-restricted fashion, to biochemically purified PDC-E2 and E3BP (which co-purify) (9/10 positive (SI > 2.76), mean SI 5.74 ± 5.04 (PDC-E2/E3BP) versus 6.67 ± 3.84 (PDC), P = NS), implying that the important PBC-specific T cell epitopes are contained within the PDC-E2 or E3BP components of PDC. Only a minority of patients responsive to PDC, however, responded to either lipoylated recombinant PDC-E2 ILD (4/10 positive, mean SI 1.98 ± 1.24, P < 0.005 versus PDC response) or lipoylated p163 (4/12 positive, mean SI 1.90 ± 1.58, P < 0.001). The lipoylation state did not affect the T cell response to either ILD or p163. Our findings suggest that in some UK patients with PBC there are immunodominant T cell autoepitopes within PDC-E2/E3BP which are outside the ILD of PDC-E2.