Figure 5.

Copurification of UGT1 with the CalS Complex.

(A) Fractionation of the CalS complex by sucrose density gradient. A Chaps-solubilized fraction of membranes from moth bean root tips was incubated with the substrate for callose synthesis. Proteins remaining associated with the product-entrapped fraction were fractionated on a sucrose density gradient (20 to 60%). CalS activity (fluorescence units [FU] per 30-μL fraction) was measured in each fraction (see Methods). CalS^HI, CalS present in high sucrose density (52%); CalS^LO, CalS present in low sucrose density (40%).

(B) Presence of UGT1 with CalS as shown by protein gel blot analysis using UGT1 antibody. Fractions from each peak were pooled, and proteins were analyzed by protein gel blotting using UGT1 antibody. Two immunoreactive bands are indicated by arrowheads (see Figure 4). CalS1 was shown to be present with these fractions (Hong et al., 2001). Sizes (in kilodaltons) of the prestained protein markers (Bio-Rad) are indicated.