Figure 2.

AlkMGO is internalized by mammalian cells, and histone AlkMGO-glycation is subjected to enzymatic regulation. HEK 293T cells were treated with increasing amounts of AlkMGO for 12 h. Following 2 h of recovery, cells were transiently transfected with (A) empty vectors or constructs to overexpress either (B) DJ-1 or (C) PAD4 for 24 h. Next, cells were harvested and fractionated to histone and soluble protein fractions. The extracted histones were derivatized with Cy5-azide as described in the Experimental Section. Samples were separated by SDS-PAGE, and gels were imaged by in gel-fluorescence followed by Coomassie Brilliant Blue staining for loading control. The soluble fractions were analyzed by western blot analysis with the indicated antibodies.