Peptide stapling is a strategy for constraining short peptides in an alpha-helical conformation. Helix stabilization by cross-linking had been shown previously to increase the helicity and potency of α-helical peptides dramatically. The stapled peptides can be used for intracellular drug targets because stapling can increase the target affinity and proteolytic resistance. The hydrocarbon-stapling is achieved by ring-closing olefin metathesis between two functionalized non-natural building blocks, either R8-S5 or S5-S5. S5 stands for S-2-(4′-pentenyl) alanine. R8 stands for R-2-(7′-octenyl) alanine. Hydrocarbon stapled α-helical peptides are capable of targeting and interfering with intracellular protein-protein interactions. These peptides contain a synthetic brace, introduced across one face of an α-helix. The structure can increase α-helical content and protease resistance, enhance target binding affinity, and promote cell membrane penetration.

      Strategy of stapled peptide synthesis is that two unnatural amino acids containing a-methyl and a-alkenyl are introduced in the process of solid phase synthesis, then the two unnatural amino acids occur olefin metathesis(RCM) Cyclization to form stable a-helix conformation of the carbon.

      1)Synthesis of unnatural amino acids containing a-methyl and a-alkenyl:

Structure of unnatural amino acids containing a-methyl and a-alkenyl

Synthesis route of unnatural amino acids containing a-methyl and a-alkenyl

      2) Synthesis of Stapled Peptide：

      Use SPPS to couple each amino acids in the sequence to peptide including two unnatural amino acids containing a-methyl and a-alkenyl; Make the cyclization using RCM Cyclization between two unnatural amino acids containing a-methyl and a-alkenyl; Fluoroarene such as biotin, FITC can be attached to peptide if necessary; Make the cleavage of peptide from resin, and we get the stapled peptide.

Synthesis Route of Stapled Peptide.