Date of Award
Doctor of Philosophy
Biochemistry, Biophysics and Molecular Biology
Understanding the molecular basis of neuroplasticity is an ambition of neuroscience. SynGAP and nNOS are two neuronal proteins which are central regulators of neuroplasticity. nNOS synthesizes nitric oxide, a second messenger important for retrograde communication between synapses, while SynGAP stimulates the GTPase activity of Ras and Rap. Regulation of these enzymes is vital for synaptic homeostasis, but there is much to learn about the details of nNOS and SynGAP activity. In this dissertation I address two questions about nNOS and SynGAP structural biology: what is the structural basis for nNOS activation by calmodulin? How do the auxiliary domains of SynGAP enable RapGAP activity?
Both nNOS and SynGAP are large proteins with modular domain organization and flexible linkers. These features make structural analysis by mainstay techniques such as NMR or X-Ray crystallography excessively difficult. Hydrogen-deuterium exchange mass spectrometry (HDX-MS) is well-suited for investigating the structure-activity relationships in proteins such as nNOS and SynGAP. Hydrogen-deuterium exchange of amide protons with the solvent, as measured in HDX-MS, provides a wealth of information on the structural dynamics of proteins. Mass spectrometry in combination with proteolytic digestion of samples enables assignment of those perturbation to discrete peptides within the protein. Using this approach, I demonstrate a role for allosteric effects in both activation of nNOS by calmodulin and the RapGAP function of SynGAP.
Hanson, Quinlin, "The role of allostery in nNOS and SynGAP activity" (2018). Graduate Theses and Dissertations. 17200.