Grant: CIHR Fellowship

The protein SOD1 can form incorrect structures in neurons. Such 'misfolding' is linked to the fatal neurodegenerative disorder ALS. SOD1 misfolding propagates within and between cells by converting correctly folded molecules, similar to infectious prions in 'mad-cow' disease. However, the molecular mechanisms of misfolding and propagation remain poorly understood, because misfolding is difficult to observe directly. We will overcome this challenge by observing the structural dynamics of single SOD1 molecules using high-resolution laser tweezers, gently tugging on the ends of a molecule to unfold and refold it repeatedly while monitoring structural changes in real time via the extension (the protein stretches under the applied force when unfolded). This approach can detect rare or transient states of SOD1-likely crucial to misfolding but difficult to observe using other means-and characterize its folding in unparalleled detail (microscopic rates, energies, existence/structural properties of intermediate/misfolded states, pathways for native folding vs. misfolding, energy landscape). To probe the mechanisms driving disease-related misfolding, we will examine the effects of mutations tied to heritable forms of ALS. The folding/misfolding of isolated mutant monomers will be measured first, before studying mutant dimers (SOD1 is normally homodimeric). Differences between the mutants and wild-type (wt) SOD1 will be examined, especially changes in the misfolded states and the pathways to misfolding (which in wt SOD1 we have shown are mediated by certain partially-native intermediates). Finally, conversion of natively folded wt SOD1 by misfolded mutant SOD1 will be observed directly by placing a misfolding-prone mutant monomer next to a wt monomer and testing changes in the wt behavior. Experiments will be amplified by computational simulations to obtain atomistic details of the misfolding. This work should help identify new targets for drugs inhibiting SOD1 misfolding.