(Figure ). This is a technique in which appropriately positioned side chain residues are linked covalently to stabilize an alpha-helical conformation of the peptide. As a model we focused on the twelve amino acid peptide ITFEDLLDYYGP-NH2 which is a ligand for the HIV capsid protein that disrupts capsid assembly in vitro, but not in cultured cells due its cell impermeability. Zhang, et al. used olefin metathesis chemistry to create a stapled version of this peptide (NYAD-1: IPFXDLLXYYGP; where X = a residue with an alkene-containing side chain) with much higher helical content than the native peptide and improved cell penetration properties. NYAD-1 is a modestly potent inhibitor of HIV replication in cultured cells. We constructed the five linear substrates shown schematically in Figure in which the amino-and Oxo-containing residues had an i-i+4 or i-i+7 spacing. The beads displaying these compounds were treated with the indicated P2CA under the standard conditions. After cleavage the products were analyzed by LC-MS. In each case, good to excellent conversion to the desired stapled peptides was observed (see Supplementary Figures ). The degree of helicity was determined by circular dichroism spectroscopy. As shown in Figure , the linear peptide was a random coil. All of the stapled MPs displayed an enhanced degree of helical content. The best of these (≈ 74% helical) was MP24, in which the i and i+4 residues had been stapled with P2CA. It is interesting that MP24, 25 and 26, which contain the same ring but were formed using a different P2CA, display markedly different levels of helicity. The two i-i+7-stapled peptides (MP27 and MP28), linked using P2CA and Py17, also displayed measurably different levels of helicity. These data suggest that the use of different P2CA units in the creation of IP +stapled MPs will provide a novel tool to fine tune the helical content of stapled peptides.