The Fl F0 ATP synthase is central to the process of cellular bioenergetics; this enzyme produces adenosine triphosphate (ATP), which is the principal form of chemical energy in the cell. In mitochondria, assembly of the Fl component of the ATP synthase is dependent on two proteins called Atp11p and Atp12p. Biochemical and genetic studies have shown that Atp11p binds the beta subunit of Fl and that Atp12p binds the alpha subunit of Fl. A principal focus of our work is to determine the molecular details that underlie Atp11p and Atp12p actions in the cell. Such efforts will be greatly facilitated by solving the crystal structure of these proteins. To this end, a purification protocol has been developed to isolate large amounts of recombinant yeast Atp11p from a bacterial expression system in a very pure form. Various conditions of pH, temperature, and additives are being explored to find the conditions to obtain crystals. Methods are also being developed to produce a soluble form of recombinant Atp12p in amounts that are sufficient to pursue crystallization trials. Concurrent with efforts to crystallize the individual Atp11p and Atp12p proteins, work is in progress to isolate complexes and co-crystallize each assembly factor with its cognate protein substrate. Determination of the three-dimensional structure of Atp11p bound to Fl beta or of Atp12p bound to Fl alpha is expected to provide important information about the mechanism of the Fl assembly factor proteins.
| Hinton, Ayana; Gatti, Domenico L; Ackerman, Sharon H (2004) The molecular chaperone, Atp12p, from Homo sapiens. In vitro studies with purified wild type and mutant (E240K) proteins. J Biol Chem 279:9016-22 |
