Fungal mitochondrial porins and DNA polymerases

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Mitochondria are membrane-bound organelles that are key to energy production in eukaryotic cells. The function of mitochondria depends on the exchange of metabolites across the two mitochondrial membranes. Our research investigates the structure of porin, a protein that allows the passage of small, water soluble molecules across the outer membrane. This molecule may also be involved in the regulation of programmed cell death, an important player in normal development, and in disease states including cancer. Therefore, an understanding of porin structure is essential in order to determine how the protein carries out these functions, and how it interacts with the rest of the cell. To this end, we are using recombinant porin from Neurospora, that is overexpressed in Escherichia coli. Different detergent systems are being evaluated as the means to fold porin in the absence of membranes, to allow structural determination by methods like nuclear magnetic resonance spectroscopy (NMR). In addition, we are investigating the signals that direct the protein to mitochondria following its synthesis, and determining the regions of the protein that interact with other proteins on both sides of the membrane, and thereby may contribute to the regulation of its function.      The second part of our research deals with mitochondrial DNA replication. Mitochondria contain small genomes that are essential to all obligately aerobic organisms. In fungi, the enzyme that replicates this DNA, the mitochondrial DNA polymerase, contains a domain of unknown function. The proposed research will investigate the function of this domain in the yeast protein, with the ultimate goal of understanding the different ways in which DNA polymerases can carry out effective, processive synthesis. Together, these projects contribute to the body of knowledge surrounding the maintenance of these essential organelles, and their interactions with the rest of the cell.