Extended Data Fig. 2: TEV(C151Dap) specifically traps its substrate in Expi293 cell lysate.
From: Mechanism-based traps enable protease and hydrolase substrate discovery
TEV variants (WT, Ala, Dap, 1 μM) were incubated with control GFP-c (a polypeptide “Gly-Gly-Gly-Ser-Gly-Gly-Gly-His6” was attached at the C-terminus of GFP) or substrate GFP-s (a TEV cleavage sequence “Glu-Asn-Leu-Tyr-Phe-Gln-Gly-His6” was attached at the C-terminus of GFP. The cleavage between Gln and Gly is underlined) in Expi293 cell lysate at 37 °C for 3 h. The concentration of GFP-s or GFP-c was (a) 1 μM or (b) 5 μM. Dap*: reaction of TEV(C151Dap) with GFP-s in Tris buffer. The input (reaction before IP) and Strep-tag enriched TEV species were analyzed by SDS-PAGE (Coomassie staining) and WB (anti-Strep for TEV and anti-GFP). (a) and (b) were repeated in three biological replicates with similar results. For gel source data, see Supplementary Fig. 1.
