Cell-free production, purification and characterization of human mitochondrial ADP/ATP carriers

Protein Expr Purif. 2018 Apr;144:46-54. doi: 10.1016/j.pep.2017.11.008. Epub 2017 Dec 5.


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.

Keywords: ADP/ATP carrier; Cell-free protein expression; Crystallization; Membrane protein; Mitochondrial carriers; Thermostability.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell-Free System
  • Gene Expression
  • Humans
  • Mitochondrial ADP, ATP Translocases / genetics
  • Mitochondrial ADP, ATP Translocases / isolation & purification*
  • Mitochondrial ADP, ATP Translocases / metabolism
  • Protein Structure, Secondary


  • Mitochondrial ADP, ATP Translocases