Figure 1.
AtSEX4 dephosphorylates glucosyl 6-P residues of both the prephosphorylated A- and B-type MDcryst. Wild-type AtSEX4 was incubated with the prelabeled A-type (A) or the B-type (B) MD in the insoluble (black symbols) or solubilized (white symbols) state using the LTA (see “Materials and Methods”). In C, two mutated AtSEX4 proteins (N333K [N→K] and K307Q [K→Q]) and the wild-type AtSEX4 protein (see insert) were incubated with the prelabeled insoluble B-type allomorph. The three phosphatases were balanced to equal p-nitrophenyl phosphate hydrolyzing activity. In A to C, prephosphorylation (and prelabeling) was performed for 60 min. At intervals, aliquots of each of the four reaction mixtures were heat treated and subjected to TLC. Radioactivity was quantified by phosphor imaging (n = 3; ±sd).