Figure 1.
Actinobacterial lysates exhibit depupylating activity that is due to Dop. (A) PanB–Pup (0.3 μM), a substrate modified with Pup using the ligase PafA, was added to cell extracts of Mycobacterium smegmatis (Msm), Corynebacterium glutamicum or Escherichia coli, and depupylation activity was monitored by anti-His6 immunoblot. (B) In Msm Δdop lysate, PanB–Pup is stable over the course of the experiment. Depupylation is restored in the complemented strain Msm Δdop–dop. Purified PanB–Pup and PanB are shown for comparison. The asterisk denotes expressed Dop-His6. Dop, deamidase of Pup; Pup, prokaryotic ubiquitin-like protein.