Methionine-Containing Peptides: Avoiding Secondary Reactions in the Final Global Deprotection

Abstract

The solid-phase synthesis of Met-containing peptides using a fluorenylmethoxycarbonyl (Fmoc)/tert-butyl (tBu) protection scheme is inevitably accompanied by two stubborn side reactions, namely, oxidation and S-alkylation (tert-butylation), which result in the formation of Met(O) and sulfonium salt impurities of the target peptide, respectively. These two reactions are acid-catalyzed, and they occur during the final trifluoroacetic (TFA)-based acidolytic cleavage step. Herein, we developed two new cleavage solutions that eradicate the oxidation and reduce S-alkylation. TFA-anisole-trimethylsilyl chloride (TMSCl)-Me2S-triisopropylsilane (TIS) containing 1 mg of triphenyl phosphine per mL of solution was the optimal mixture for Cys-containing peptides, while for the remaining peptides, TIS was not required. Both cleavage solutions proved to be excellent when sensitive amino acids such as Cys and Trp were involved. TMSCl did not affect either of these sensitive amino acids. Reversing the sulfonium salt to free Met-containing peptide was achieved by heating the peptide at 40 °C for 24 h using 5% acetic acid.