FIGURE 4.

AdoMet cross-linking studies of the human SET1 family members. a, subset of SET1 family members undergoing auto-methylation. Isolated SET domains were incubated with [³H]AdoMet in the presence (+) or absence (−) of 250 μm unmodified H3 peptide. The upper panel shows a Coomassie Blue-stained SDS-polyacrylamide gel, and the lower panel shows [³H]methyl incorporation by fluorography after a 24-h exposure to film. b, UV exposure was used to cross-link [³H]AdoMet to isolated SET1 family SET domains (upper panels) or SET1 family core complexes (lower panels). All panels represent [³H]methyl incorporation by fluorography after a 3-day exposure to film. A single asparagine to alanine mutation in the AdoMet binding pocket of each SET domain abolishes [³H]AdoMet cross-linking. c, quantification of [³H]AdoMet cross-linking in the presence and absence of WRAD. Bands corresponding to the SET domains of each wild type SET1 family member were excised and quantified by liquid scintillation counting as described under “Experimental Procedures.” Error bars represent the S.E. of measurement from three independent experiments.