TY - JOUR AU - Szabó, Eszter AU - Nemes-Nikodém, Éva AU - Vass, Krisztina Rubina AU - Zámbó, Zsófia Melinda AU - Zrupko, E. AU - Törőcsik, Beáta AU - Ozohanics, Olivér AU - Nagy, Bálint AU - Ambrus, Attila TI - Structural and Biochemical Investigation of Selected Pathogenic Mutants of the Human Dihydrolipoamide Dehydrogenase JF - INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES J2 - INT J MOL SCI VL - 24 PY - 2023 IS - 13 PG - 23 SN - 1661-6596 DO - 10.3390/ijms241310826 UR - https://m2.mtmt.hu/api/publication/34075046 ID - 34075046 N1 - Export Date: 1 September 2023 Correspondence Address: Ambrus, A.; Department of Biochemistry, 37-47 Tuzolto St, Hungary; email: ambrus.attila@med.semmelweis-univ.hu Funding details: TKP2021-EGA-25, ÚNKP-19-3-III-SE-17 Funding details: Horizon 2020 Framework Programme, H2020, PID18322 Funding details: Helmholtz-Zentrum Berlin für Materialien und Energie, HZB Funding details: Semmelweis Egyetem, STIA-OTKA-2021 Funding details: Hungarian Scientific Research Fund, OTKA, 143627 Funding details: Helmholtz-Zentrum für Umweltforschung, UFZ Funding details: Nemzeti Kutatási, Fejlesztési és Innovaciós Alap, NKFIA Funding text 1: This research was funded by the Hungarian Brain Research Program 2 (2017-1.2.1-NKP-2017-00002 grant, to Vera Adam-Vizi), Semmelweis University (STIA-OTKA-2021 grant, to A.A.), Hungarian Scientific Research Fund (OTKA grant 143627, to A.A.), Ministry of Innovation and Technology of Hungary (TKP2021-EGA-25 grant, to A.A. and ÚNKP-19-3-III-SE-17 grant of the New National Excellence Program, to E.S.; project no. TKP2021-EGA-25 has been implemented with the support provided by the Ministry of Innovation and Technology of Hungary from the National Research, Development and Innovation Fund, financed under the TKP2021-EGA funding scheme), and European Union’s Horizon 2020 Research and Innovation Programme (Structural Biology Research Infrastructures for Translational Research and Discovery, iNEXT-Discovery PID18322 and MX-212-00269-ST (Helmholtz Zentrum Berlin/CALIPSO) grants, both to A.A.). Funding text 2: The authors gratefully acknowledge the generous support from Vera Adam-Vizi (Semmelweis University) and Manfred S. Weiss (Helmholtz-Zentrum Berlin). AB - Clinically relevant disease-causing variants of the human dihydrolipoamide dehydrogenase (hLADH, hE3), a common component of the mitochondrial α-keto acid dehydrogenase complexes, were characterized using a multipronged approach to unravel the molecular pathomechanisms that underlie hLADH deficiency. The G101del and M326V substitutions both reduced the protein stability and triggered the disassembly of the functional/obligate hLADH homodimer and significant FAD losses, which altogether eventually manifested in a virtually undetectable catalytic activity in both cases. The I12T-hLADH variant proved also to be quite unstable, but managed to retain the dimeric enzyme form; the LADH activity, both in the forward and reverse catalytic directions and the affinity for the prosthetic group FAD were both significantly compromised. None of the above three variants lent themselves to an in-depth structural analysis via X-ray crystallography due to inherent protein instability. Crystal structures at 2.89 and 2.44 Å resolutions were determined for the I318T- and I358T-hLADH variants, respectively; structure analysis revealed minor conformational perturbations, which correlated well with the residual LADH activities, in both cases. For the dimer interface variants G426E-, I445M-, and R447G-hLADH, enzyme activities and FAD loss were determined and compared against the previously published structural data. © 2023 by the authors. LA - English DB - MTMT ER - TY - JOUR AU - Tőkés, Anna-Mária AU - Vári-Kakas, S. AU - Kulka, Janina AU - Törőcsik, Beáta TI - Tumor Glucose and Fatty Acid Metabolism in the Context of Anthracycline and Taxane-Based (Neo)Adjuvant Chemotherapy in Breast Carcinomas JF - FRONTIERS IN ONCOLOGY J2 - FRONT ONCOL VL - 12 PY - 2022 PG - 23 SN - 2234-943X DO - 10.3389/fonc.2022.850401 UR - https://m2.mtmt.hu/api/publication/32804574 ID - 32804574 LA - English DB - MTMT ER - TY - JOUR AU - Mihályi, Csaba AU - Iordanov, Iordan AU - Törőcsik, Beáta AU - Csanády, László TI - Simple binding of protein kinase A, prior to phosphorylation, allows CFTR anion channels to be opened by nucleotides JF - PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA J2 - P NATL ACAD SCI USA VL - 117 PY - 2020 IS - 35 SP - 21740 EP - 21746 PG - 7 SN - 0027-8424 DO - 10.1073/pnas.2007910117 UR - https://m2.mtmt.hu/api/publication/31397038 ID - 31397038 LA - English DB - MTMT ER - TY - JOUR AU - Szabó, Eszter AU - Wilk, Piotr AU - Nagy, Bálint AU - Zámbó, Zsófia Melinda AU - Bui, Dávid AU - Weichsel, Andrzej AU - Arjunan, Palaniappa AU - Törőcsik, Beáta AU - Hubert, Agnes AU - Furey, William AU - Montfort, William R AU - Jordan, Frank AU - Weiss, Manfred S AU - Ádám, Veronika AU - Ambrus, Attila TI - Underlying molecular alterations in human dihydrolipoamide dehydrogenase deficiency revealed by structural analyses of disease-causing enzyme variants JF - HUMAN MOLECULAR GENETICS J2 - HUM MOL GENET VL - 28 PY - 2019 IS - 20 SP - 3339 EP - 3354 PG - 16 SN - 0964-6906 DO - 10.1093/hmg/ddz177 UR - https://m2.mtmt.hu/api/publication/30799456 ID - 30799456 AB - Human dihydrolipoamide dehydrogenase (hLADH, hE3) deficiency (OMIM# 246900) is an often prematurely lethal genetic disease usually caused by inactive or partially inactive hE3 variants. Here we report the crystal structure of wild-type hE3 at an unprecedented high resolution of 1.75 Å and the structures of six disease-causing hE3 variants at resolutions ranging from 1.44 to 2.34 Å. P453L proved to be the most deleterious substitution in structure as aberrations extensively compromised the active site. The most prevalent G194C-hE3 variant primarily exhibited structural alterations close to the substitution site, whereas the nearby cofactor-binding residues were left unperturbed. The G426E substitution mainly interfered with the local charge distribution introducing dynamics to the substitution site in the dimer interface; G194C and G426E both led to minor structural changes. The R460G, R447G, and I445M substitutions all perturbed a solvent accessible channel, the so-called H+/H2O channel, leading to the active site. Molecular pathomechanisms of enhanced reactive oxygen species (ROS) generation and impaired binding to multienzyme complexes were also addressed according to the structural data for the relevant mutations. In summary, we present here for the first time a comprehensive study that links three-dimensional structures of disease-causing hE3 variants to residual hLADH activities, altered capacities for ROS generation, compromised affinities for multienzyme complexes, and eventually clinical symptoms. Our results may serve as useful starting points for future therapeutic intervention approaches. LA - English DB - MTMT ER - TY - JOUR AU - Csanády, László AU - Törőcsik, Beáta TI - Cystic fibrosis drug ivacaftor stimulates CFTR channels at picomolar concentrations JF - ELIFE J2 - ELIFE VL - 8 PY - 2019 PG - 18 SN - 2050-084X DO - 10.7554/eLife.46450 UR - https://m2.mtmt.hu/api/publication/30775333 ID - 30775333 AB - The devastating inherited disease cystic fibrosis (CF) is caused by mutations of the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) anion channel. The recent approval of the CFTR potentiator drug ivacaftor (Vx-770) for the treatment of CF patients has marked the advent of causative CF therapy. Currently, thousands of patients are being treated with the drug, and its molecular mechanism of action is under intensive investigation. Here we determine the solubility profile and true stimulatory potency of Vx-770 towards wild-type (WT) and mutant human CFTR channels in cell-free patches of membrane. We find that its aqueous solubility is ~200 fold lower (~60 nanomolar), whereas the potency of its stimulatory effect is >100 fold higher, than reported, and is unexpectedly fully reversible. Strong, but greatly delayed, channel activation by picomolar Vx-770 identifies multiple sequential slow steps in the activation pathway. These findings provide solid guidelines for the design of in vitro studies using Vx-770. © 2019, Csanády and Töröcsik. LA - English DB - MTMT ER - TY - JOUR AU - Zahola, Péter AU - Hanics, János AU - Pintér, Panka AU - Máté, Zoltán AU - Gáspárdy, Anna AU - Hevesi, Zsófia AU - Echevarria, Diego AU - Adori, Csaba AU - Barde, Swapnali AU - Törőcsik, Beáta AU - Erdélyi, Ferenc AU - Szabó, Gábor AU - Wagner, Ludwig AU - Kovács, Gábor Géza AU - Hökfelt, Tomas AU - Harkany, Tibor AU - Alpár, Alán TI - Secretagogin expression in the vertebrate brainstem with focus on the noradrenergic system and implications for Alzheimer's disease JF - BRAIN STRUCTURE & FUNCTION J2 - BRAIN STRUCT FUNC VL - 224 PY - 2019 IS - 6 SP - 2061 EP - 2078 PG - 18 SN - 1863-2653 DO - 10.1007/s00429-019-01886-w UR - https://m2.mtmt.hu/api/publication/30723780 ID - 30723780 N1 - SE NAP B Research Group of Experimental Neuroanatomy and Developmental Biology, Semmelweis University, Budapest, Hungary Department of Anatomy, Semmelweis University, Budapest, Hungary Institute of Experimental Medicine, Hungarian Academy of Sciences, Budapest, Hungary Department of Molecular Neurosciences, Center for Brain Research, Medical University of Vienna, Vienna, 1090, Austria Institute of Neuroscience, University of Miguel Hernandez de Elche, Alicante, Spain Department of Neuroscience, Karolinska Institutet, Biomedicum 7D, Stockholm, SE-17165, Sweden Department of Medical Biochemistry, Semmelweis University, Budapest, Hungary Department of Internal Medicine III, Medical University of Vienna, Vienna, Austria Institute of Neurology, Medical University of Vienna, Vienna, Austria Cited By :12 Export Date: 31 October 2023 Correspondence Address: Alpár, A.; SE NAP B Research Group of Experimental Neuroanatomy and Developmental Biology, Hungary; email: Alpar.Alan@med.semmelweis-univ.hu AB - Calcium-binding proteins are widely used to distinguish neuronal subsets in the brain. This study focuses on secretagogin, an EF-hand calcium sensor, to identify distinct neuronal populations in the brainstem of several vertebrate species. By using neural tube whole mounts of mouse embryos, we show that secretagogin is already expressed during the early ontogeny of brainstem noradrenaline cells. In adults, secretagogin-expressing neurons typically populate relay centres of special senses and vegetative regulatory centres of the medulla oblongata, pons and midbrain. Notably, secretagogin expression overlapped with the brainstem column of noradrenergic cell bodies, including the locus coeruleus (A6) and the A1, A5 and A7 fields. Secretagogin expression in avian, mouse, rat and human samples showed quasi-equivalent patterns, suggesting conservation throughout vertebrate phylogeny. We found reduced secretagogin expression in locus coeruleus from subjects with Alzheimer's disease, and this reduction paralleled the loss of tyrosine hydroxylase, the enzyme rate limiting noradrenaline synthesis. Residual secretagogin immunoreactivity was confined to small submembrane domains associated with initial aberrant tau phosphorylation. In conclusion, we provide evidence that secretagogin is a useful marker to distinguish neuronal subsets in the brainstem, conserved throughout several species, and its altered expression may reflect cellular dysfunction of locus coeruleus neurons in Alzheimer's disease. LA - English DB - MTMT ER - TY - JOUR AU - Szabó, Eszter AU - Mizsei, Réka AU - Wilk, P AU - Zámbó, Zsófia Melinda AU - Törőcsik, Beáta AU - Weiss, MS AU - Ádám, Veronika AU - Ambrus, Attila TI - Crystal structures of the disease-causing D444V mutant and the relevant wild type human dihydrolipoamide dehydrogenase JF - FREE RADICAL BIOLOGY AND MEDICINE J2 - FREE RADICAL BIO MED VL - 124 PY - 2018 SP - 214 EP - 220 PG - 7 SN - 0891-5849 DO - 10.1016/j.freeradbiomed.2018.06.008 UR - https://m2.mtmt.hu/api/publication/3407990 ID - 3407990 N1 - Department of Medical Biochemistry, MTA-SE Laboratory for Neurobiochemistry, Semmelweis University, Budapest, H-1094, Hungary Macromolecular Crystallography, Helmholtz-Zentrum Berlin für Materialien und Energie, D-12489, Berlin, Germany Cited By :2 Export Date: 24 January 2020 CODEN: FRBME Correspondence Address: Ambrus, A.; Department of Medical Biochemistry, Semmelweis University, 37-47 Tuzolto Street, Hungary; email: ambrus.attila@med.semmelweis-univ.hu Chemicals/CAS: dihydrolipoamide dehydrogenase, 37340-89-9, 9001-18-7; Dihydrolipoamide Dehydrogenase; Mutant Proteins Funding details: Hungarian Scientific Research Fund Funding details: Hungarian Scientific Research Fund, 112230 Funding details: Magyar Tudományos Akadémia, 02001 Funding text 1: Financial support was secured from the Hungarian Academy of Sciences (MTA grant 02001 to V.A-V.), the Hungarian Scientific Research Fund (OTKA grant 112230 to V.A-V.), the Hungarian Brain Research Program (grants KTIA_13_NAP-A-III/6. and 2017-1.2.1-NKP-2017-00002 to V.A-V.), the EMBO , Fulbright , and Bolyai Fellowships (to A.A.), and the Young Investigator Research Grants of the Semmelweis University and Gedeon Richter Pharmaceuticals PIc . (to A.A.). Data collection at Helmholtz-Zentrum Berlin was carried out under contract number 16204087-ST. Department of Medical Biochemistry, MTA-SE Laboratory for Neurobiochemistry, Semmelweis University, Budapest, H-1094, Hungary Macromolecular Crystallography, Helmholtz-Zentrum Berlin für Materialien und Energie, D-12489, Berlin, Germany Cited By :3 Export Date: 4 January 2021 CODEN: FRBME Correspondence Address: Ambrus, A.; Department of Medical Biochemistry, Semmelweis University, 37-47 Tuzolto Street, Hungary; email: ambrus.attila@med.semmelweis-univ.hu Chemicals/CAS: dihydrolipoamide dehydrogenase, 37340-89-9, 9001-18-7; Dihydrolipoamide Dehydrogenase; Mutant Proteins Funding details: Hungarian Scientific Research Fund, OTKA Funding details: Magyar Tudományos Akadémia, MTA Funding details: Hungarian Scientific Research Fund, OTKA, 112230 Funding details: Magyar Tudományos Akadémia, MTA, 02001 Funding text 1: Financial support was secured from the Hungarian Academy of Sciences (MTA grant 02001 to V.A-V.), the Hungarian Scientific Research Fund (OTKA grant 112230 to V.A-V.), the Hungarian Brain Research Program (grants KTIA_13_NAP-A-III/6. and 2017-1.2.1-NKP-2017-00002 to V.A-V.), the EMBO , Fulbright , and Bolyai Fellowships (to A.A.), and the Young Investigator Research Grants of the Semmelweis University and Gedeon Richter Pharmaceuticals PIc . (to A.A.). Data collection at Helmholtz-Zentrum Berlin was carried out under contract number 16204087-ST. Department of Medical Biochemistry, MTA-SE Laboratory for Neurobiochemistry, Semmelweis University, Budapest, H-1094, Hungary Macromolecular Crystallography, Helmholtz-Zentrum Berlin für Materialien und Energie, D-12489, Berlin, Germany Cited By :3 Export Date: 5 January 2021 CODEN: FRBME Correspondence Address: Ambrus, A.; Department of Medical Biochemistry, Semmelweis University, 37-47 Tuzolto Street, Hungary; email: ambrus.attila@med.semmelweis-univ.hu Chemicals/CAS: dihydrolipoamide dehydrogenase, 37340-89-9, 9001-18-7; Dihydrolipoamide Dehydrogenase; Mutant Proteins Funding details: Hungarian Scientific Research Fund, OTKA Funding details: Magyar Tudományos Akadémia, MTA Funding details: Hungarian Scientific Research Fund, OTKA, 112230 Funding details: Magyar Tudományos Akadémia, MTA, 02001 Funding text 1: Financial support was secured from the Hungarian Academy of Sciences (MTA grant 02001 to V.A-V.), the Hungarian Scientific Research Fund (OTKA grant 112230 to V.A-V.), the Hungarian Brain Research Program (grants KTIA_13_NAP-A-III/6. and 2017-1.2.1-NKP-2017-00002 to V.A-V.), the EMBO , Fulbright , and Bolyai Fellowships (to A.A.), and the Young Investigator Research Grants of the Semmelweis University and Gedeon Richter Pharmaceuticals PIc . (to A.A.). Data collection at Helmholtz-Zentrum Berlin was carried out under contract number 16204087-ST. AB - We report the crystal structures of the human (dihydro)lipoamide dehydrogenase (hLADH, hE3) and its disease-causing homodimer interface mutant D444V-hE3 at 2.27 and 1.84 Å resolution, respectively. The wild type structure is a unique uncomplexed, unliganded hE3 structure with the true canonical sequence. Based on the structural information a novel molecular pathomechanism is proposed for the impaired catalytic activity and enhanced capacity for reactive oxygen species generation of the pathogenic mutant. The mechanistic model involves a previously much ignored solvent accessible channel leading to the active site that might be perturbed also by other disease-causing homodimer interface substitutions of this enzyme. © 2018 Elsevier Inc. LA - English DB - MTMT ER - TY - JOUR AU - Sorum, Ben AU - Törőcsik, Beáta AU - Csanády, László TI - Asymmetry of movements in CFTR's two ATP sites during pore opening serves their distinct functions JF - ELIFE J2 - ELIFE VL - 6 PY - 2017 PG - 17 SN - 2050-084X DO - 10.7554/eLife.29013 UR - https://m2.mtmt.hu/api/publication/3287727 ID - 3287727 AB - CFTR, the chloride channel mutated in cystic fibrosis (CF) patients, is opened by ATP binding to two cytosolic nucleotide binding domains (NBDs), but pore-domain mutations may also impair gating. ATP-bound NBDs dimerize occluding two nucleotides at interfacial binding sites; one site hydrolyzes ATP, the other is inactive. The pore opens upon tightening, and closes upon disengagement, of the catalytic site following ATP hydrolysis. Extent, timing, and role of non-catalytic-site movements are unknown. Here we exploit equilibrium gating of a hydrolysis-deficient mutant and apply Phi value analysis to compare timing of opening-associated movements at multiple locations, from the cytoplasmic ATP sites to the extracellular surface. Marked asynchrony of motion in the two ATP sites reveals their distinct roles in channel gating. The results clarify the molecular mechanisms of functional cross-talk between canonical and degenerate ATP sites in asymmetric ABC proteins, and of the gating defects caused by two common CF mutations. LA - English DB - MTMT ER - TY - JOUR AU - Szabó, Eszter AU - Mizsei, Réka AU - Zámbó, Zsófia Melinda AU - Törőcsik, Beáta AU - Weiss, Manfred S AU - Ádám, Veronika AU - Ambrus, Attila TI - Crystal structure of the D444V disease-causing mutant of human dihydrolipoamide dehydrogenase JF - BIOCHIMICA ET BIOPHYSICA ACTA-BIOENERGETICS J2 - BBA-BIOENERGETICS VL - 1857 PY - 2016 IS - Suppl. SP - e100 SN - 0005-2728 DO - 10.1016/j.bbabio.2016.04.337 UR - https://m2.mtmt.hu/api/publication/3262509 ID - 3262509 LA - English DB - MTMT ER - TY - JOUR AU - Ambrus, Attila AU - Wang, J AU - Mizsei, Réka AU - Zámbó, Zsófia Melinda AU - Törőcsik, Beáta AU - Jordan, F AU - Ádám, Veronika TI - Structural alterations induced by ten disease-causing mutations of human dihydrolipoamide dehydrogenase analyzed by hydrogen/deuterium-exchange mass spectrometry: Implications for the structural basis of E3 deficiency JF - BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR BASIS OF DISEASE J2 - BBA-MOL BASIS DIS VL - 1862 PY - 2016 IS - 11 SP - 2098 EP - 2109 PG - 12 SN - 0925-4439 DO - 10.1016/j.bbadis.2016.08.013 UR - https://m2.mtmt.hu/api/publication/3114546 ID - 3114546 N1 - Attila Ambrus, Junjie Wang and Reka Mizsei contributed equally to this work. Department of Medical Biochemistry, MTA-SE Laboratory for Neurobiochemistry, Semmelweis University, Budapest, Hungary Department of Chemistry, Rutgers University, Newark, NJ, United States Laboratory of Mass Spectrometry and Gaseous Ion Chemistry, The Rockefeller University, New York, NY, United States Laboratory of Immunobiology, Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA, United States Cited By :9 Export Date: 28 May 2020 CODEN: BBADE Correspondence Address: Ambrus, A.; Department of Medical Biochemistry, MTA-SE Laboratory for Neurobiochemistry, Semmelweis UniversityHungary; email: ambrus.attila@med.semmelweis-univ.hu Chemicals/CAS: amino acid, 65072-01-7; deuterium, 7782-39-0; dihydrolipoamide dehydrogenase, 37340-89-9, 9001-18-7; hydrogen, 12385-13-6, 1333-74-0 Funding details: Magyar Tudományos Akadémia, MTA Funding details: Hungarian Scientific Research Fund, OTKA Funding details: Hungarian Scientific Research Fund, OTKA, 112230 Funding details: Magyar Tudományos Akadémia, MTA, 02001 Funding text 1: We are grateful to Drs. Oliver Ozohanics, Karoly Vekey (both from the Research Centre for Natural Sciences, Hungarian Academy of Sciences, Budapest, Hungary) and Arpad Somogyi (Ohio State University, Columbus, OH, USA) for their contributions to the method development in mass spectrometry. Financial support is gratefully acknowledged from the Hungarian Academy of Sciences (MTA grant 02001 to V.A.-V.), the Hungarian Scientific Research Fund (OTKA, grant 112230 to V.A.-V.), the Hungarian Brain Research Program (grant KTIA_13_NAP-A-III/6 to V.A.-V.), the Bolyai and the Fulbright Fellowships (to A.A.), NIH - R15GM116077 and NSF-CHE - 1402675 (to F.J.). Department of Medical Biochemistry, MTA-SE Laboratory for Neurobiochemistry, Semmelweis University, Budapest, Hungary Department of Chemistry, Rutgers University, Newark, NJ, United States Laboratory of Mass Spectrometry and Gaseous Ion Chemistry, The Rockefeller University, New York, NY, United States Laboratory of Immunobiology, Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA, United States Cited By :12 Export Date: 5 January 2021 CODEN: BBADE Correspondence Address: Ambrus, A.; Department of Medical Biochemistry, MTA-SE Laboratory for Neurobiochemistry, Semmelweis UniversityHungary; email: ambrus.attila@med.semmelweis-univ.hu Chemicals/CAS: amino acid, 65072-01-7; deuterium, 7782-39-0; dihydrolipoamide dehydrogenase, 37340-89-9, 9001-18-7; hydrogen, 12385-13-6, 1333-74-0 Funding details: Hungarian Scientific Research Fund, OTKA Funding details: Magyar Tudományos Akadémia, MTA Funding details: Hungarian Scientific Research Fund, OTKA, 112230 Funding details: Magyar Tudományos Akadémia, MTA, 02001 Funding text 1: We are grateful to Drs. Oliver Ozohanics, Karoly Vekey (both from the Research Centre for Natural Sciences, Hungarian Academy of Sciences, Budapest, Hungary) and Arpad Somogyi (Ohio State University, Columbus, OH, USA) for their contributions to the method development in mass spectrometry. Financial support is gratefully acknowledged from the Hungarian Academy of Sciences (MTA grant 02001 to V.A.-V.), the Hungarian Scientific Research Fund (OTKA, grant 112230 to V.A.-V.), the Hungarian Brain Research Program (grant KTIA_13_NAP-A-III/6 to V.A.-V.), the Bolyai and the Fulbright Fellowships (to A.A.), NIH - R15GM116077 and NSF-CHE - 1402675 (to F.J.). AB - Pathogenic amino acid substitutions of the common E3 component (hE3) of the human alpha-ketoglutarate dehydrogenase and the pyruvate dehydrogenase complexes lead to severe metabolic diseases (E3 deficiency), which usually manifest themselves in cardiological and/or neurological symptoms and often cause premature death. To date, 14 disease-causing amino acid substitutions of the hE3 component have been reported in the clinical literature. None of the pathogenic protein variants has lent itself to high-resolution structure elucidation by X-ray or NMR. Hence, the structural alterations of the hE3 protein caused by the disease-causing mutations and leading to dysfunction, including the enhanced generation of reactive oxygen species by selected disease-causing variants, could only be speculated. Here we report results of an examination of the effects on the protein structure of ten pathogenic mutations of hE3 using hydrogen/deuterium-exchange mass spectrometry (HDX-MS), a new and state-of-the-art approach of solution structure elucidation. On the basis of the results, putative structural and mechanistic conclusions were drawn regarding the molecular pathogenesis of each disease-causing hE3 mutation addressed in this study. © 2016 Elsevier B.V. LA - English DB - MTMT ER -