On the effect of alkaline pH and cofactor availability in the conformational and oligomeric state of Escherichia coli glutamate decarboxylase
| Autor: | D Verzili, Malcolm Buckle, C. Mérigoux, Fabio Giovannercole, Gaia Grassini, C Zamparelli, P Vachette, Daniela De Biase |
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| Přispěvatelé: | Institut de Biologie Intégrative de la Cellule (I2BC), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS), Fonction et Architecture des Assemblages Macromoléculaires (FAAM), Département Biochimie, Biophysique et Biologie Structurale (B3S), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Institut de Biologie Intégrative de la Cellule (I2BC), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS), Laboratoire de biologie et pharmacologie appliquée (LBPA), Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Ecole Normale Supérieure Paris-Saclay (ENS Paris Saclay), Institut de biochimie et biophysique moléculaire et cellulaire (IBBMC), Université Paris-Sud - Paris 11 (UP11)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Biologie et de Pharmacologie Appliquée (LBPA), École normale supérieure - Cachan (ENS Cachan)-Centre National de la Recherche Scientifique (CNRS) |
| Jazyk: | angličtina |
| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
analytic ultracentrifugation glutamate decarboxylase limited proteolysis pyridoxal 5′-phosphate (PLP) small angle X-ray scattering (SAXS) Decarboxylation Proteolysis [SDV]Life Sciences [q-bio] 030106 microbiology Glutamate decarboxylase Bioengineering medicine.disease_cause Biochemistry Cofactor 03 medical and health sciences chemistry.chemical_compound X-Ray Diffraction Escherichia coli medicine [SDV.BBM]Life Sciences [q-bio]/Biochemistry Molecular Biology Protein Structure Quaternary Molecular Biology Pyridoxal chemistry.chemical_classification medicine.diagnostic_test biology Escherichia coli Proteins Membrane Proteins Hydrogen-Ion Concentration Enzyme Catalytic cycle chemistry biology.protein Protein Multimerization Biotechnology |
| Zdroj: | Protein Engineering, Design and Selection Protein Engineering, Design and Selection, Oxford University Press (OUP), 2017, 30, pp.237-246. ⟨10.1093/protein/gzw076⟩ Protein Engineering, Design and Selection, Oxford University Press (OUP), 2016, 30, pp.237-246. ⟨10.1093/protein/gzw076⟩ Protein Engineering, Design and Selection, 2017, 30, pp.237-246. ⟨10.1093/protein/gzw076⟩ |
| ISSN: | 1741-0126 1741-0134 |
| Popis: | International audience; Escherichia coli glutamate decarboxylase (EcGad) is a homohexameric pyridoxal 5′-phosphate (PLP)-dependent enzyme. It is the structural component of the major acid resistance system that protects E. coli from strong acid stress (pH < 3), typically encountered in the mammalian gastro-intestinal tract. In fact EcGad consumes one proton/catalytic cycle while yielding γ-aminobutyrate and carbon dioxide from the decarboxylation of L-glutamate. Two isoforms of Gad occur in E. coli (GadA and GadB) that are 99% identical in sequence. GadB is the most intensively investigated. Prompted by the observation that some transcriptomic and proteomic studies show EcGad to be expressed in conditions far from acidic, we investigated the structural organization of EcGadB in solution in the pH range 7.5-8.6. Small angle X-ray scattering, combined with size exclusion chromatography , and analytical ultracentrifugation analysis show that the compact and entangled EcGadB hexameric structure undergoes dissociation into dimers as pH alkalinizes. When PLP is not present, the dimeric species is the most abundant in solution, though evidence for the occurrence of a likely tetrameric species was also obtained. Trp fluorescence emission spectra as well as limited proteolysis studies suggest that PLP plays a key role in the acquisition of a folding necessary for the canonical catalytic activity. |
| Databáze: | OpenAIRE |
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