The metal loaded SOD1 protein is an enormously stable dimer. However, it has recently been shown that dissociation of SOD1 can occur under quite mild conditions through modification of two of its structural elements. Disulfide bond reduction between residues C57 and C146 in the metal-free form of SOD1 results in complete dissociation of the protein to its monomeric subunits at physiologically relevant pH [81,111,132]. However, the disulfide-reduced, metal-bound protein remains a stable homodimer. The binding of a single zinc ion shifts monomeric SOD1 back to its dimeric form . Taking into account that approximately one-third of SOD1 may be apo for metal ions and that the environment of the cell is reducing, there could exist a significant pool of SOD1 monomer in vivo. Monomeric SOD1 also likely plays a functional role, particularly in the context of copper insertion by CCS, where the heterodimer model of copper incorporation into SOD1 requires one monomer of apo SOD1 to form a complex with one monomer of CCS . There is some evidence that copper transfer from yeast CCS to yeast SOD1 may be mediated through an intermolecular disulfide involving Cys57 of SOD1. Thus, a reduced disulfide bond for SOD1 recognition and metal loading by CCS is required . The disulfide bond is proposed to be formed concomitant with copper ion transfer from yeast CCS to yeast SOD1 resulting in the mature form of the enzyme .
How might the fALS mutations affect the quaternary structure of SOD1? fALS mutants of the MBR class (Figure 4, right subunit) have decreased affinity for metals in comparison to the wild-type protein and are thus more prone to dissociate, depending on the status of the intrasubunit disulfide bond . In contrast, some WTL mutations (black spheres in Figure 1B) may directly destabilize dimeric SOD1 by perturbing the dimer interface. These same mutations may indirectly destablize the protein through metal deficiency if they interfere with CCS/SOD1 complex formation and metal transfer to SOD1. Other WTL mutations in the ^-barrel (medium gray spheres in Figure 1B) may destabilize the ^-barrel fold, slowing the maturation process. We recently discovered, using analytical ultracentrifugation, that purified fALS SOD1 mutants A4V and G93A in their metal-free form are partially monomeric, even without reduction of the disulfide bond . These mutants also demonstrate an increased susceptibility for reduction of their disulfide bond in comparison to the wild-type protein . Finally, the immature form of SOD1 is more prone to oxidative damage, underscoring the role that this form may play in the disease state .
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