Figure 6. ncOGT is sufficient for O-GlcNAcylation of mitochondrial proteins.
(A) HEK 293 suspension cells were transfected with either GFP or GFP-tagged ncOGT. Mitochondria, purified from transfected cells, were incubated with buffer or with 10 μg/ml of proteinase K. Lysates from the mitochondrial or the cytosolic fractions were probed with anti-OGT, anti-α-tubulin (cytosol), anti-ATPB (mitochondrial matrix) and anti-TOM20 (mitochondrial outer membrane) antibodies. (B) O-GlcNAc levels in mitochondrial fractions were detected with anti-O-GlcNAc antibody (CTD 110.6). The amount of protein loaded in each lane in (A) is 10 μg, whereas approximately 100 μg was used in (B).