FIG 6.

Multiturnover ubiquitylation reactions with SCF demonstrate that the presence of R54D ubiquitin diminishes the lengths of the polyubiquitin chains on product compared with WT ubiquitin and that the introduction of D143K Ube2R2 with R54D ubiquitin substantially increases the lengths of the chains. Data represent time courses for the ubiquitylation of β-catenin (β-Cat) peptide for the entire double mutant cycle. Notice that some of the polyubiquitin chains in the reaction mixture containing WT Ube2R2 and ubiquitin migrated as high-molecular-weight smears, and while these were still present in the reaction mixtures containing D143K Ube2R2, the intensities had shifted significantly lower in the direction of the substrate band.