Mutations in the active site Cys residue (C94) abolish the activity of SCE, but some still allow loading with SUMO (see the text). Mutations in the first alpha helix (Lys residues 15, 19 or 28) have more impact on SUMO chain formation than on the mono-sumoylation activity of SCE. In contrast, mutation of Lys residues of the carboxyl-terminal region (Lys residues 146, 147, 150 and 154) to Arg has little impact on any activity of the enzyme.