Gain in functions of mutant Cu,Zn-superoxide dismutases as a causative factor in familial amyotrophic lateral sclerosis: Less reactive oxidant formation but high spontaneous aggregation and precipitation
A. Okado-matsumoto et al., Gain in functions of mutant Cu,Zn-superoxide dismutases as a causative factor in familial amyotrophic lateral sclerosis: Less reactive oxidant formation but high spontaneous aggregation and precipitation, FREE RAD RE, 33(1), 2000, pp. 65-73
Eight mutant Cu,Zn-superoxide dismutases (SODs) related to familial amyotro
phic lateral sclerosis (FALS) were produced in a baculovirus/insect cell ex
pression system and their molecular properties in terms of hydroxyl radical
formation and aggregation were compared with the wild-type enzyme. Treatme
nt of the enzymes with Chelex 100 resin decreased Cu contents as well as SO
D activities in all mutant Cu,Zn-SODs, indicating that the affinities of th
e Enzymes for copper ion were decreased. Contrary to previous reports, all
the mutant Cu,Zn-SODs exhibited less reactive oxidant producing ability in
the presence of hydrogen peroxide than the wild-type enzyme. Both SOD activ
ities and their reactive oxidant forming correlated well with the copper io
n content of the molecules. In addition, the proteins spontaneously aggrega
ted and were precipitated by simple centrifugation at 12,000g for 20 min in
keeping their enzyme activities. Since hyaline inclusions found in FALS pa
tients with SOD1 mutations contained components which were reactive to anti
-Cu,Zn-SOD antibody, a primary reaction caused by mutant SOD1 slay be attri
buted to their propensity to form aggregates. Aggregated but still active m
utant SOD1 would be expected to mediate the formation of reactive oxygen sp
ecies and nitrosylation in a more condensed state.