Cyclic Modular β-Sheets

The development of peptide β-hairpins is problematic, because folding depends on the amino acid sequence and changes to the sequence can significantly decrease folding. Robust β-hairpins that can tolerate such changes are attractive tools for studying interactions involving protein β-sheets and developing inhibitors of these interactions. This paper introduces a new class of peptide models of protein β-sheets that addresses the problem of separating folding from the sequence. These model β-sheets are macrocyclic peptides that fold in water to present a pentapeptide β-strand along one edge; the other edge contains the tripeptide β-strand mimic Hao [JACS 2000, 122, 7654] and two additional amino acids. The pentapeptide and Hao-containing peptide strands are connected by two δ-linked ornithine (δOrn) turns [JACS 2003, 125, 876]. Each δOrn turn contains a free α-amino group that permits the linking of individual modules to form divalent β-sheets. These “cyclic modular β-sheets” are synthesized by standard solid-phase peptide synthesis of a linear precursor followed by solution-phase cyclization. Eight cyclic modular β-sheets 1a − 1h containing sequences based on β-amyloid and macrophage inflammatory protein 2 were synthesized and characterized by 1H NMR. Linked cyclic modular β-sheet 2, which contains two modules of 1b, was also synthesized and characterized. 1H NMR studies show downfield α-proton chemical shifts, δOrn δ-proton magnetic anisotropy, and NOE cross-peaks that establish all compounds but 1c and 1g to be moderately or well folded into a conformation that resembles a β-sheet. Pulsed-field gradient NMR diffusion experiments show little or no self-association at low (≤2 mM) concentrations. Changes to the residues in the Hao-containing strands of 1c and 1g improve folding and show that folding of the structures can be enhanced without altering the sequence of the pentapeptide strand. Well-folded cyclic modular β-sheets 1a, 1b, and 1f each have a phenylalanine directly across from Hao, suggesting that cyclic modular β-sheets containing aromatic residues across from Hao are better folded.