The H+ -ATPase of mitochondria, choloroplasts, and bacteria synthesizes ATP during respiration, providing the principal energy source for the cell. An important question in the field of Bioenergetics concerns the mechanism of this vitally important enzyme. The interactions within, and between, the F1 and F0 components that constitute the enzyme complex play an intimate role in the energy transduction reactions. The studies described on ATPase biogenesis address the fundamental question of how functionally important subunit contacts are initiated. The long-term goal of this project is to identify, and to define the function of, nuclear gene products required for biosynthesis of the mitochondrial F1-F0 complex in the yeast Saccharomyces cerevisiae. The basic objective is to provide information on the structure of the mitochondrial ATPase and to determine how the structure influences the function of the enzyme. The project is also aimed at answering the more global question of how macromolecular protein complexes are assembled from their component subunits. The present focus is specifically on assembly of the F1 component of the enzyme. Previous work with yeast mutants defective in F1 assembly has revealed a new class of proteins which are required for assembly of the F1 oligomer. An immediate goal is to purify these proteins and to use them in studies that will define their roles in F1 assembly. Extragenic revertants have already been isolated from yeast with mutations in the assembly factor genes. The suppressors will be cloned to identify proteins that interact directly with the F1 assembly factors. Strains are also available which are defective in F1 assembly due to mutations in the alpha and beta structural subunit genes. These mutants will be used to obtain extragenic revertants. The acquisition of these revertants will be important for two reasons. First, this will identify which of the alpha and beta subunit mutants are affected in domains that interact directly with other proteins. Second, identification of the suppressors subunits into will disclose proteins that are required for assembly of the alpha and beta subunits into the F1 oligomer.
