Exploration of incorporation of N -methylated amino acids into peptides by sense-suppression method

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Here we describe the ribosomal synthesis of peptides containing an N(alpha)-methylated amino acid by using sense-suppression method. To incorporate the various N(alpha)-methylated amino acids into the peptide, we utilized recently developed two technologies, a ribozyme-based tRNA acylation system (Flexizyme system) and a reconstituted cell-free translation system (PURE system). Firstly, various N(alpha)-methylated amino acids were charged onto a suppressor tRNA by Flexizyme system. Then, using PURE system, we constructed a vacant codon corresponding to the suppressed position. By combining these systems, we were able to examine various N(alpha)-methylated amino acids for the single incorporation into the peptide backbone and determined those with high efficiencies. Thus, this demonstration opens a new door to develop a new methodology for the synthesis of peptides containing multiple N-methylated backbone via the genetic code reprogramming.