Figure 4.

Shift in the electrophoretic mobility of tissue transglutaminases after binding to nucleotides. Purified TG isolated from human erythrocytes (A and B) and from guinea pig liver (C) were examined by electrophoresis in agarose (3% in A and B; 2% in C) with (lanes 2, 3, and 4) and without (lane 1) admixture (45 min at room temperature) of nucleotides/Mg²⁺ (0.4 mM of GMP, GDP, or GTP). The proteins were prepared at concentrations of 3.3 μM in a buffer of 75 mM imidazole-HCl, pH 7.2, containing 0.5 mM EDTA. Samples of 30 μl (i.e., 8 μg protein per lane) were applied to the gel, and electrophoresis was carried out for 2 hr at 4°C in the above buffer. The gels with light backgrounds in the picture were developed with Coomassie blue; the segment shown in B Right, probing the electrophoretic mobility of KCl-stripped red cell TG, was photographed under UV light after a transamidase activity-specific staining procedure, based on the incorporation of fluorescent dansylcadaverine into dimethylated casein (29, 30).