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Protein aggregation and phase separation appear to play important roles in diseases like amyotrophic lateral sclerosis (ALS) and frontotemporal lobar dementia (FTLD), but the interplay between different participating molecular events-which may facilitate or inhibit one another-can be difficult to study by conventional ensemble methods. In a recent study, Kevin Rhine and co-workers make use of point mutations to demonstrate the contrasting behaviour of condensates arising from Glycine and Arginine FUS mutants using single molecules fluorescence measurements. [ABSTRACT FROM AUTHOR]

The misfolding and mutation of Cu/Zn superoxide dismutase (SOD1) is commonly associated with amyotrophic lateral sclerosis (ALS). SOD1 can accumulate within stress granules (SGs), a type of membraneless organelle, which is believed to form via liquid–liquid phase separation (LLPS). Using wild‐type, metal‐deficient, and different ALS disease mutants of SOD1 and computer simulations, we report here that the absence of Zn leads to structural disorder within two loop regions of SOD1, triggering SOD1 LLPS and amyloid formation. The addition of exogenous Zn to either metal‐free SOD1 or to the severe ALS mutation I113T leads to the stabilization of the loops and impairs SOD1 LLPS and aggregation. Moreover, partial Zn‐mediated inhibition of LLPS was observed for another severe ALS mutant, G85R, which shows perturbed Zn‐binding. By contrast, the ALS mutant G37R, which shows reduced Cu‐binding, does not undergo LLPS. In addition, SOD1 condensates induced by Zn‐depletion exhibit greater cellular toxicity than aggregates formed by prolonged incubation under aggregating conditions. Overall, our work establishes a role for Zn‐dependent modulation of SOD1 conformation and LLPS properties that may contribute to amyloid formation. Synopsis: The formation of membraneless organelles, such as stress granules (SGs), is believed to occur through the process of liquid–liquid phase separation (LLPS). The amyotrophic lateral sclerosis (ALS) disease‐associated protein SOD1 is known to accumulate within SGs, but if it itself undergoes LLPS, and if this affects amyloid formation, is not well‐studied. The apo form of SOD1 undergoes LLPS.SOD1 LLPS and subsequent aggregation are regulated by a conformational transition between a disordered and a relatively compact folded state.The metal cofactor Zn prevents SOD1 LLPS via a conformational transition.SOD1 condensates show greater cytotoxicity than aggregates. [ABSTRACT FROM AUTHOR]

Aggregation of Cu–Zn superoxide dismutase (SOD1) is implicated in the motor neuron disease, amyotrophic lateral sclerosis (ALS). Although more than 140 disease mutations of SOD1 are available, their stability or aggregation behaviors in membrane environment are not correlated with disease pathophysiology. Here, we use multiple mutational variants of SOD1 to show that the absence of Zn, and not Cu, significantly impacts membrane attachment of SOD1 through two loop regions facilitating aggregation driven by lipid-induced conformational changes. These loop regions influence both the primary (through Cu intake) and the gain of function (through aggregation) of SOD1 presumably through a shared conformational landscape. Combining experimental and theoretical frameworks using representative ALS disease mutants, we develop a ‘co-factor derived membrane association model’ wherein mutational stress closer to the Zn (but not to the Cu) pocket is responsible for membrane association-mediated toxic aggregation and survival time scale after ALS diagnosis. [ABSTRACT FROM AUTHOR]

Superoxide dismutase (SOD) is the primary enzyme of the cellular antioxidant defense cascade. Misfolding, concomitant oligomerization, and higher order aggregation of human cytosolic SOD are linked to amyotrophic lateral sclerosis (ALS). Although, with two metal ion cofactors SOD1 is extremely robust, the de-metallated apo form is intrinsically disordered. Since the rise of oxygen-based metabolism and antioxidant defense systems are evolutionary coupled, SOD is an interesting protein with a deep evolutionary history. We deployed statistical analysis of sequence space to decode evolutionarily co-varying residues in this protein. These were validated by applying graph theoretical modelling to understand the impact of the presence of metal ion co-factors in dictating the disordered (apo) to hidden disordered (wild-type SOD1) transition. Contact maps were generated for different variants, and the selected significant residues were mapped on separate structure networks. Sequence space analysis coupled with structure networks helped us to map the evolutionarily coupled co-varying patches in the SOD1 and its metal-depleted variants. In addition, using structure network analysis, the residues with a major impact on the internal dynamics of the protein structure were investigated. Our results reveal that the bulk of these evolutionarily co-varying residues are localized in the loop regions and positioned differentially depending upon the metal residence and concomitant steric restrictions of the loops. [ABSTRACT FROM AUTHOR]