This application describes an approach to better understand the structure of the interaction between the Krebs/Tricarboxylic acid (TCA) cycle and the propionate pathway. Then it uses enzymatic assays to probe the function of proteins at the intersection of these pathways. Finally it examines samples from subjects with propionate disorders to better characterize this structure and function. The Problem: Although Krebs/TCA cycle was identified greater than 80 years ago and the propionate pathway disorders were identified greater than 40 years ago, their structural interaction has not been described. Individuals with propionate disorders have long term complications which implicate energy production and this interaction between propionate pathway and Krebs/TCA cycle may provide a site of therapy. By understanding how these pathways intersect-we suspect as an physical association-- and assay them as a whole system, we are looking for possible sites for therapeutics. The Approach: To characterize this interaction, we aim to isolate the association formed at the intersection of these pathways. We then will probe this structure for function by using existing assays to determine if they can be used as screening assays. Finally we aim to see how the structure and function differs in cells from individuals/mice with the propionate pathway disorders, propionic acidemia and methylmalonic aciduria. Anticipated Results: We have preliminary data that shows the Krebs/TCA cycle enzymes, succinate synthase and oxoglutarate dehydrogenase complex, interact with the propionate pathway enzyme, methylmalonyl CoA as a complex. We anticipate that we will be able to isolate the association involving the propionate pathway and Krebs/TCA cycle enzymes and it will be associated with inner mitochondrial membrane.
We aim to isolate the interaction and be able to assay it using pre-existing methods designed for individual enzymes. These assays and isolation of a complex will inform us of possible sites for therapeutics and a method for screening small molecules. Finally, we will be able to assay tissues and cells from individuals and mice with propionate pathway disorders and be able to identify differences and potentially new sites for therapeutics for these disorders.
We suspect that the Krebs/tricarboxylic acid cycle and propionate pathway, two energy producing pathways that intersect, interact as an association. Here we propose identifying components of this association and then assaying the components as an association. Identifying the components of this association and screening the association for enzyme activity provides a mechanism to screen new treatments for propionate pathway disorders such as propionic acidemia and methylmalonic aciduria, as well as other disorders which affect the Krebs/TCA cycle's function.
