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. Author manuscript; available in PMC: 2014 Sep 18.
Published in final edited form as: Bioconjug Chem. 2013 Aug 22;24(9):10.1021/bc4002618. doi: 10.1021/bc4002618

Figure 6.

Figure 6

The overall intein-based release and functionalization method is performed in three steps, as shown in this example for an scFv. Step 1: Yeast displaying scFv-intein fusion proteins are reacted with MESNA for 45 min. Step 2: Released proteins in the MESNA solution (a mixture of scFv-thioesters and scFv-intein-Aga2p fusions) are removed from the yeast and cysteine alkyne is added. This reaction proceeds for 20 h to complete release of the scFv from intein and to conjugate the alkyne via EPL. Step 3: The alkyne-conjugated scFvs are reacted in a CuAAC reaction with biotin alkyne for 1 h or with an azido surface for 16 h.