Developing short peptides into useful probes and therapeutic leads remains a difficult challenge. Structural rigidification is a proven method for improving the properties of short peptides. In this work, we report a strategy for stabilizing peptide macrocycles by introducing side-chain-to-side-chain staples, producing peptide bicycles with higher affinity, selectivity, and resistance to degradation. We have applied this strategy to G1, an 11-residue peptide macrocycle that binds the Src homology 2 (SH2) domain of growth-factor-bound protein 2 (Grb2). Several homodetic peptide bicycles were synthesized entirely on-resin with high yields. Two rounds of iterative design produced peptide bicycle BC1, which is 60-fold more potent than G1 and 200-fold more selective. Also, BC1 is completely intact after 24 hours in buffered human serum, conditions under which G1 is completely degraded. Our peptide bicycle approach holds promise for the development of selective inhibitors of SH2 domains and other pTyr-binding proteins, as well as inhibitors of many other protein-protein interactions.

While peptides are promising as probes and therapeutics, targeting intracellular proteins will require greater understanding of highly structured, cell-internalized scaffolds. We recently reported BC1, an 11-residue bicyclic peptide that inhibits the Src homology 2 (SH2) domain of growth factor receptor-bound protein 2 (Grb2). In this work, we describe the unique structural and cell uptake properties of BC1 and similar cyclic and bicyclic scaffolds. These constrained scaffolds are taken up by mammalian cells despite their net neutral or negative charges, while unconstrained analogs are not. The mechanism of uptake is shown to be energy-dependent and endocytic, but distinct from that of Tat. The solution structure of BC1 was investigated by NMR and MD simulations, which revealed discrete water-binding sites on BC1 that reduce exposure of backbone amides to bulk water. This represents an original and potentially general strategy for promoting cell uptake.

Novel nickel(II) complexes of pyridine-azamacrocycles (PyMACs) with pendant arms have been synthesized using simple, direct, and selective mono-N-functionalization of PyMACs. These complexes have been characterized by spectroscopy and X-ray crystallography. Nickel(II)-PyMAC complexes with a flexible pendant arm bearing a tertiary amine, a carboxylic acid, or an amide group exhibit structural and color changes due to "on-off" arm coordination to the metal center. Five- or six-coordinate complexes with the arm bound to the nickel(II) center are high-spin, while their four-coordinate "arm-off" counterparts are low-spin. Synergistic axial coordination of acetonitrile and the amide group from the pendant arm was observed. Coordination to the nickel(II) center lowers the pK(a) of the functional group attached to the macrocycle via a propylene linker by up to 4-5 orders of magnitude. Varying hydrogen bonding and proton-donating properties of the pendant arm affects the peroxidase-like activity of Ni(II)-PyMAC complexes in the oxidation of ABTS with hydrogen peroxide.

The inside cover picture shows a new strategy for designing constrained peptides that involves careful introduction of side‐chain‐to‐side chain crosslinks within a head‐to‐tail peptide macrocycle. For details of how it was used to produce peptide bicycles that target Grb2 with higher potency, better selectivity, and greater resistance to degradation, see the communication by J. A. Kritzer et al. on