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Zinc-release assay: The release of zinc, in the presence of a weak chelator (PAR), from the H46R/H48Q Sod1 mutant (DM) was measured spectroscopically in comparison with wild type (WT) Sod1. The DM cannot bind copper in the active site and leaches zinc upon the addition of PAR, which is exacerbated by the reducing agent TCEP. Ccs1 binding by the disulfide reduced DM Sod1 molecule stabilizes zinc binding and prohibits chelation by PAR.
