Dynamic properties of the G93A mutant of copper-zinc superoxide dismutase as detected by NMR spectroscopy: Implications for the pathology of familial amyotrophic lateral sclerosis

The backbone assignment of the copper-zinc superoxide dismutase amyotrophic lateral sclerosis G93A mutant was performed on an N-15-enriched protein sample. N-15 R-1, R-2, and R-1p and N-15-H-1 NOE experiments were then carried out at 600 MHz on G93A Cu2Zn2SOD and the values compared to the dynamics data for the wild-type protein. In addition, N-15 and H-1 chemical shift comparisons between wild-type Cu2Zn2SOD and its G93A mutant were also made. G93A exhibits a higher mobility than wildtype Cu2Zn2SOD, particularly in loops III and V, on a time scale faster than the rate of protein tumbling. There are also distinct chemical shift and NOE differences in residues 35-42 and 92-95, which comprise these loops. These two regions Of Cu2Zn2SOD form the end of the beta-barrel termed the beta-barrel plug [Tainer, J. A., Getzoff, E. D., Beem, K. M., Richardson, J. S., and Richardson, D. C. (1982) J. Mol. Biol. 160, 181-217]. The increased mobility and reduction of the number of observed NOES in this region indicate an opening of the beta-barrel that may lead to amyloid fibrillogenesis. Alternatively, a motor neuron-specific substrate may bind this region of the protein, leading to deleterious modifications and/or reactions.