1. Cell-free production, purification and characterization of human mitochondrial ADP/ATP carriers
- Author
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Delphine Baud, Stéphanie Ravaud, Céline Juillan-Binard, Aleksandra Woznicka-Misaila, Eva Pebay-Peyroula, Institut de biologie structurale (IBS - UMR 5075 ), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), and Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)
- Subjects
0301 basic medicine ,Gene isoform ,Gene Expression ,Mitochondrion ,Cell-free protein expression ,Protein Structure, Secondary ,03 medical and health sciences ,Mitochondrial carriers ,Humans ,Protein secondary structure ,Mitochondrial ADP-ATP Carriers ,Thermostability ,Cell-Free System ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,Chemistry ,Yeast ,030104 developmental biology ,Membrane ,Membrane protein ,Biochemistry ,ADP/ATP carrier ,Crystallization ,Mitochondrial ADP, ATP Translocases ,Biotechnology - Abstract
International audience; Mitochondrial Carriers (MCs) are responsible for fluent traffic of a variety of compounds that need to be shuttled via mitochondrial inner membranes to maintain cell metabolism. The ADP/ATP Carriers (AACs) are responsible for the import of ADP inside the mitochondria and the export of newly synthesized ATP. In human, four different AACs isoforms are described which are expressed in tissue-specific manner. They are involved in different genetic diseases and play a role in cancerogenesis. Up to now only the structures of the bovine (isoform 1) and yeast (isoforms 2 and 3) AAC have been determined in one particular conformation, obtained in complex with the CATR inhibitor. Herein, we report that full-length human ADP/ATP Carriers isoform 1 and 3 were successfully expressed in cell-free system and purified in milligram amounts in detergent-solubilized state. The proteins exhibited the expected secondary structure content. Thermostability profiles showing stabilization by the CATR inhibitor suggest that the carriers are well folded.
- Published
- 2018
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