title: A dual conformation of the post-decarboxylation intermediate is associated with distinct enzyme states in mycobacterial KGD (α-ketoglutarate decarboxylase).
abstract: α-Ketoacid dehydrogenases are large multi-enzyme machineries that orchestrate the oxidative decarboxylation of α-ketoacids with the concomitant production of acyl-CoA and NADH. The first reaction, catalysed by α-ketoacid decarboxylases (E1 enzymes), needs a thiamine diphosphate cofactor and represents the overall rate-limiting step. Although the catalytic cycles of E1 from the pyruvate dehydrogenase (E1p) and branched-chain α-ketoacid dehydrogenase (E1b) complexes have been elucidated, little structural information is available on E1o, the first component of the α-ketoglutarate dehydrogenase complex, despite the central role of this complex at the branching point between the TCA (tricarboxylic acid) cycle and glutamate metabolism. In the present study, we provide structural evidence that MsKGD, the E1o (α-ketoglutarate decarboxylase) from Mycobacterium smegmatis, shows two conformations of the post-decarboxylation intermediate, each one associated with a distinct enzyme state. We also provide an overall picture of the catalytic cycle, reconstructed by either crystallographic snapshots or modelling. The results of the present study show that the conformational change leading the enzyme from the initial (early) to the late state, although not required for decarboxylation, plays an essential role in catalysis and possibly in the regulation of mycobacterial E1o.
authors.affiliation: *Institut Pasteur, Unité de Microbiologie Structurale and CNRS UMR3528, 25 rue du Docteur Roux, 75724 Paris Cedex 15, France.
authors.forename: Tristan
authors.initials: T
authors.lastname: Wagner
authors.affiliation: *Institut Pasteur, Unité de Microbiologie Structurale and CNRS UMR3528, 25 rue du Docteur Roux, 75724 Paris Cedex 15, France.
authors.forename: Nathalie
authors.initials: N
authors.lastname: Barilone
authors.affiliation: *Institut Pasteur, Unité de Microbiologie Structurale and CNRS UMR3528, 25 rue du Docteur Roux, 75724 Paris Cedex 15, France.
authors.forename: Pedro M
authors.initials: PM
authors.lastname: Alzari
authors.affiliation: *Institut Pasteur, Unité de Microbiologie Structurale and CNRS UMR3528, 25 rue du Docteur Roux, 75724 Paris Cedex 15, France.
authors.forename: Marco
authors.initials: M
authors.lastname: Bellinzoni
mesh_terms: D000272:Adipates / Q000737:chemistry / Q000378:metabolism; D019880:Aldehyde-Ketone Transferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D055162:Biocatalysis; D002262:Carboxy-Lyases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D020134:Catalytic Domain; D003653:Decarboxylation; D007655:Ketoglutarate Dehydrogenase Complex / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007656:Ketoglutaric Acids / Q000737:chemistry / Q000378:metabolism; D008958:Models, Molecular; D008968:Molecular Conformation; D062105:Molecular Docking Simulation; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D020102:Mycobacterium smegmatis / Q000201:enzymology*; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D058849:Protein Refolding; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
chemical_list: D000272:Adipates; D001426:Bacterial Proteins; D007656:Ketoglutaric Acids; D050505:Mutant Proteins; D010446:Peptide Fragments; D011994:Recombinant Proteins; C001918:alpha-ketoadipic acid; D007655:Ketoglutarate Dehydrogenase Complex; D019880:Aldehyde-Ketone Transferases; C021414:2-hydroxy-3-oxoadipate synthase; D002262:Carboxy-Lyases; C021474:2-oxoglutarate decarboxylase