Presentation description

Cdc48 (also called VCP and p97) is an essential protein and multifunctional ATPase found primarily in lower eukaryotes. Cdc48 is involved in protein degradation, metabolic regulation, and organelle and cell recycling. It functions primarily by binding to other, smaller proteins known as adapters, allowing it to tailor its enzymatic activity to specific use cases. In a study done by a former Shen Lab member, a previously uncharacterized interaction between Cdc48 and the enzyme CAR2 was observed, indicating the possibility of CAR2 functioning as a Cdc48 adapter. CAR2 (Carbonic anhydrase 2) is a small enzyme involved in nitrogen metabolism and arginine degradation. It has been implicated in numerous diseases, including pancreatic adenocarcinoma, osteopetrosis, and renal tubular acidosis. Understanding the nature of the interaction between Cdc48 and CAR2 will help to understand the many metabolic processes in which both proteins are involved. As such, we set out to characterize the effect of Cdc48 on CAR2 activity, as well as the effect of CAR2 on Cdc48 activity. To do this, we used various methodologies, including adapting a CAR2 enzymatic activity assay based on NADH oxidation, performing a malachite green ATPase assay to observe Cdc48 activity, as well as using mass photometry and SDS-PAGE silver stain gels for protein quality control and quantification. Based on these methods, our preliminary data suggests that Cdc48 increases CAR2 activity, while the presence of CAR2 decreases Cdc48 activity. While these trends are interesting, they are preliminary and require additional data. Our next steps will focus on collecting more data and repeating experiments to further confirm these findings to conclude the potential interaction of Cdc48 and CAR2.