Fig. 4. Creation of the first ‘lanthiprotein’ through minimal stapling. (a) Protein lanthionine formation from MEK1-C218C222 via regioselective generation of Dha222 and intramolecular reaction with nearby Cys218. Prior studies in MEK1 suggested32 a wide spectrum of reagent activities from the panel (Fig. 2c) that might allow differential installation of Dha.$Mobs. = 46 057.¶Mobs. = 45 627 (plus other side products, Mobs. = 45 658, 18% plus 45 951, 59%). (b) Formation of lanthionine MEK1-Lan218222 from MEK1-Cys218–Cys222 with DIB Mcalc. = 45 658, Mobs. = 45 663. Treatment of the product with β-mercaptoethanol (BME) and the addition of DBDGla suggested that neither Dha222 nor Cys218 were present in the mono-desulfurized product. (c) MSMS of tryptic digest peptides confirmed the formation of lanthionine; observation of Lan in MEK206–227 shown. (d) MS1 chromatogram suggested a DL/LL – Lan epimeric mixture.
