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. 2018 Sep 17;293(46):17754–17768. doi: 10.1074/jbc.RA118.004185

Figure 8.

Figure 8.

OGT Swap mutant displays Rp-αS-UDP-GlcNAc diastereomer co-substrate sensitivity for E10S glycosylation. A, in vitro glycosylation of HCF3R-ASA substrate was assayed using either no OGT (lane 1) or WT (lane 2) or mutant Swap (lane 3) or K842M (lane 4) OGT. Substrate glycosylation was detected using RL2 anti-O-GlcNAc antibody. HCF3R-ASA protein (IB) was detected with anti-His antibody. B, in vitro glycosylation of HCF3R-ASA was assayed using either no OGT (lane 1) or WT or Swap OGT with 0.5 mm UDP-GlcNAc (lanes 2 and 3), Sp-αS-UDP-GlcNAc (lanes 4 and 5), or Rp-αS-UDP-GlcNAc (lanes 6 and 7). Substrate glycosylation was detected using RL2 anti-O-GlcNAc antibody. Anti-His antibody (IB) was used to detect the HCF3R-ASA protein. C, in vitro cleavage assay of HCF3R-AEA. The assay conditions are as in B. Anti-His antibody was used to detect uncleaved and cleaved HCF3R-ASA proteins. Black rectangle, uncleaved HCF3R substrate; black circle, cleaved product.