Cells expressing the S2168 mutant alleles (indicated on top) were harvested and treated with the membrane-impermeant thiol reagent MTSES at the time indicated below each lane, subjected to organic denaturation and delipidation, PEGylated with PEG-maleimide, and analyzed by SDS-PAGE and western blotting, as described in Materials and Methods. The asterisk, “+1 PEG”, and “+2 PEG” indicate the position of the unmodified monomer, the singly-PEGylated and the doubly-PEGylated species, respectively.