FIGURE 4.

H46R and H48Q form none or few HMW complexes compared with other ALS-linked SOD1 mutants. Fresh NSC34 cell lysates containing WT or various SOD1-FLAG mutants were incubated with the cross-linker DSG for 60 min before the reaction was quenched. The reaction mixtures were resolved in 10% SDS-PAGE and blotted with a FLAG antibody. After cross-linking, WT SOD1 migrated as both monomer and dimer; however, all mutants with the exception of H46R and H48Q showed formation of HMW complexes in addition to monomeric and dimeric forms. As the control in SDS-PAGE, all SOD1-FLAG ran as monomers in the absence of DSG (second panel), and actin blotting showed equal loading of all samples (third panel).