De novo design and synthesis of an ice-binding, dendrimeric, polypeptide based on insect antifreeze proteins

A new strategy is presented for the design and synthesis of peptides that exhibit ice-binding and antifreeze activity. A pennant-type dendrimer polypeptide scaffold combining an α-helical backbone with four short β-strand branches was synthesized in solid phase using Fmoc chemistry in a divergent approach. The 51-residue dendrimer was characterized by reverse phase high performance liquid chromatography, mass spectrometry and circular dichroism. Each β-strand branch contained three overlapping TxT amino acid repeats, an ice-binding motif found in the ice-binding face of the spruce budworm (Choristoneura fumiferana) and beetle (Tenebrio molitor) antifreeze proteins. Ice crystals in the presence of the polypeptide monomer displayed f at, hexagonal plate morphology, similar to that produced by weakly active antifreeze proteins. An oxidized dimeric form of the dendrimer polypeptide also produced f at hexagonal ice crystals and was capable of inhibiting ice crystal growth upon temperature reduction, a phenomenon termed thermal hysteresis, a def ning property of antifreeze proteins. Linkage of the pennant-type dendrimer to a tri-functional cascade-type polypeptide produced a trimeric macromolecule that gave f at hexagonal ice crystals with higher thermal hysteresis activity than the dimer or monomer and an ice crystal.