Date of Graduation
Master of Science in Chemistry (MS)
Chemistry & Biochemistry
Second Committee Member
Pure sciences; Biological sciences; Protein; Ras; Tuberous sclerosis
The Ras superfamily of G-proteins are of great research interest for structure-function relationships among proteins as they act as molecular switches in the regulation of various biochemical reactions in the cell. They are regulated by protein-protein interactions targeted to the highly flexible switch regions. Mutations in G-proteins or their effectors may cause alterations in structure and/or function that can lead to overactivity.
The Ras-related protein Cell division cycle 42 (Cdc42) is important in regulating cell-signaling processes. The T35A mutation in Cdc42 leads to a decrease in flexibility of the Switch I region responsible for effector binding. The kinetics of the intrinsic GTP hydrolysis reaction were compared for Cdc42 (WT and T35A) in the absence and presence of a peptide derivative of PAK21 activated kinase (PBD46). The mutation does not affect the intrinsic GTP hydrolysis rate of Cdc42, but binding with PBD46 is altered by the mutation; in the presence of PBD46, GTP hydrolysis is completely inhibited for Cdc42 WT while partial recovery of GTP hydrolysis for Cdc42 T35A was observed in the presence of PBD46. The data obtained lead us to propose two slowly interconverting conformational states of Cdc42 T35A, both able to hydrolyze GTP, but one that shows a weakened affinity for PBD46.
Ras homolog enriched in brain (Rheb) is a member of the Ras superfamily of proteins known to regulate cell growth and proliferation by signaling the mammalian target of rapamycin (mTOR) pathway. The Tuberous Sclerosis Complex 2 protein (TSC2) regulates Rheb by functioning as a GTPase activating protein (GAP). At present, very little is known about the molecular features of the Rheb-TSC2 protein interaction. We present biochemical and biophysical data on the interaction of Rheb with TSC2 WT, using a shorter, 218 amino acid GTPase derivative of TSC2 (TSC2-218), as well as with the mutant TSC2-218 K114A, to characterize the TSC2-218 constructs and their GAP activity towards Rheb. A decrease in secondary structure as well as a loss of GAP activity is seen in the K114A mutant compared to WT.
Morris, Kyla Marie Morinini, "Biochemical and Biophysical Studies of Novel Features of Ras-related Protein Interactions" (2013). Theses and Dissertations. 1002.