Structural characterization of zinc-deficient human superoxide dismutase and implications for ALS

Area of Science:

• Biochemistry
• Neuroscience
• Structural Biology

Background:

• Over 130 mutations in copper, zinc superoxide dismutase (SOD) are linked to familial amyotrophic lateral sclerosis (ALS), causing selective motor neuron death in 25% of patients.
• ALS-associated SOD mutations are hypothesized to induce structural defects and misfolding, decreasing the enzyme's affinity for zinc.
• Loss of zinc from SOD is sufficient to trigger apoptosis in motor neurons in vitro, highlighting the critical role of the zinc site.

Purpose of the Study:

• To investigate the structural and pathogenic significance of the zinc-binding site in human SOD.
• To elucidate the structural consequences of zinc deficiency in SOD using a designed zinc-deficient mutant.

Main Methods:

• Determined the 2.0-A-resolution crystal structure of a designed zinc-deficient human SOD mutant.
• The mutant was engineered by replacing two zinc-binding ligands with serine residues.

Main Results:

• The crystal structure revealed a significant 9-degree twist between SOD subunits, widening the dimer interface and representing the largest observed intersubunit rotational shift.
• Key structural alterations included partial disorder in the electrostatic and zinc-binding loops, rotation of the catalytic Arg143 residue away from the active site, and dual conformations for the Cys57-Cys146 disulfide bridge.
• Zinc deficiency increased SOD's redox activity by allowing greater access to catalytic copper and destabilized the dimer interface, leading to increased aggregation with thiol reductants. However, heterodimerization with wild-type Cu,Zn SOD prevented aggregation.

Conclusions:

• The structural instability induced by zinc deficiency in SOD, including subunit rotation and altered active site conformation, provides insights into the neuropathology of ALS.
• The observed destabilization of the dimer interface and increased lability of the disulfide bridge in zinc-deficient SOD contribute to its pathogenic potential.
• Rapid heterodimerization of zinc-deficient SOD with wild-type Cu,Zn SOD may explain the dominant inheritance pattern observed in familial ALS cases linked to SOD mutations.