文摘
Mutations in copper-zinc superoxide dismutase (CuZnSOD) cause 25% of familial amyotrophiclateral sclerosis (FALS) cases. This paper examines one such mutant, H46R, which has no superoxidedismutase activity yet presumably retains the gain-of-function activity that leads to disease. We demonstratethat Cu2+ does not bind to the copper-specific catalytic site of H46R CuZnSOD and that Cu2+ competeswith other metals for the zinc binding site. Most importantly, Cu2+ was found to bind strongly to a surfaceresidue near the dimer interface of H46R CuZnSOD. Cysteine was identified as the new binding site onthe basis of multiple criteria including UV-vis spectroscopy, RR spectroscopy, and chemical derivatization.Cysteine 111 was pinpointed as the position of the reactive ligand by tryptic digestion of the modifiedprotein and by mutational analysis. This solvent-exposed residue may play a role in the toxicity of thisand other FALS CuZnSOD mutations. Furthermore, we propose that the two cysteine 111 residues, foundon opposing subunits of the same dimeric enzyme, may provide a docking location for initial metal insertionduring biosynthesis of wild-type CuZnSOD in vivo.