PROTEIN FOLD SWITCHING: INVESTIGATING THE MECHANISM OF αβ-PLAIT TO 3α FOLD INTERCONVERSION

Loading...
Thumbnail Image

Files

Solomon_umd_0117E_20482.pdf (24.42 MB)
(RESTRICTED ACCESS)
No. of downloads:

Publication or External Link

Date

2019

Citation

Abstract

Naturally occurring metamorphic proteins have the ability to interconvert from one folded state to another through either a limited set of mutations or by way of a change in the local environment. However, the design of these types of proteins has remained challenging. This dissertation shows that it is possible to switch reversibly between two different but common folds employing only temperature changes. The study demonstrates that a latent 3α state can be unmasked from an αβ-plait topology with a single V90T amino acid substitution in a designed system, populating both forms simultaneously. The equilibrium between these two states exhibits temperature dependence, such that the 3α state is predominant (>90%) at 5°C, while the αβ-plait fold is the major species (>90%) at 30°C. The structure and dynamics of these two temperature-dependent topologies, as well as their energetics and kinetics of interconversion, are characterized utilizing NMR spectroscopy. Additional analysis show that the temperature-dependent characteristics of the 3α<->αβ-plait fold switch can be modulated by mutations. Stability studies through H-D exchange approach provide insight on the energetic basis for temperature induced 3α<->αβ-plait fold conversion. Further investigations demonstrated that interconversion between the 3α and αβ-plait states can be triggered by additional environmental factors including pressure, ligand binding, and redox state. This dissertation adds to the growing body of literature on protein fold metamorphism providing the first description of switching between two distinct monomeric protein folds using only temperature or pressure. Additionally, the studies of ligand- and redox-induced 3α<->αβ-plait fold switching emphasize the ability to mimic by design some of the mechanisms of fold interconversion that are found in naturally occurring metamorphic proteins. Given the high occurrence of the 3α and αβ-plait folds in the universe of known protein structures, the results suggest that such fold switching events may have occurred in the evolutionary expansion of function for natural versions of these topologies.

Notes

Rights