DE-NOVO DESIGN AND STRUCTURAL CHARACTERIZATION OF AN ALPHA-HELICAL HAIRPIN PEPTIDE - A MODEL SYSTEM FOR THE STUDY OF PROTEIN-FOLDING INTERMEDIATES

The de novo design and structural characterization of an alpha-helical hairpin peptide (alpha-helix/turn/alpha-helix, alpha t alpha) are rep orted. The peptide is intended to provide a model system for the study of the interactions of secondary structural elements during protein f olding. Both the diffusion-collision and framework models of protein f olding envision that the earliest intermediates in protein folding are transient secondary structures or microdomains which interact and bec ome mutually stabilizing. Design principles for the alpha t alpha pept ide were drawn from the large body of work on the structure of peptide s in solution. Computer modeling was not used in the design process. S tudy of alpha t alpha by circular dichroism and two-dimensional nuclea r magnetic resonance indicates that the designed peptide is monomeric, helical, and stable in aqueous solution at room temperature. Analysis of two-dimensional nuclear magnetic resonance experiments indicates t hat the two helices and the turn form in the intended positions and th at the helices associate in the designed orientation. Development of a lpha t alpha represents an advance in protein design in that both the secondary structural elements and designed tertiary interactions have been realized and can be detected in solution by nuclear magnetic reso nance. The resulting model system resembles a protein folding intermed iate and will allow the study of interacting helices in a context that approximates an early stage in protein folding.