NMR STRUCTURE OF A DE-NOVO DESIGNED, PEPTIDE 33MER WITH 2 DISTINCT, COMPACT BETA-SHEET FOLDS

A de novo designed 33-residue polypeptide folds as a compact beta-shee t sandwich tetramer in aqueous solution. NMR structural analysis shows that although monomer subunits have the same three-stranded antiparal lel beta-sheet fold, two equally populated conformational states are i dentified. Conformational heterogeneity arises from formation of two d istinct dimer folds. Each dimer is formed by continuing the monomer be ta-sheet into a six-stranded sheet similar to that found in alpha-chem okines. Dimer heterogeneity arises primarily from a two-residue shift in the alignment of interfacial strands. NOE-based conformational mode ling has yielded well-defined structures for both dimer types. While t he tetramer beta-sheet sandwich most probably results from association of hydrophobic surfaces from two amphipathic dimers, dimers could com bine to form either two types of homotetramers and/or one heterotetram er composed of both dimer types. Even though interdimer NOEs could not be unambiguously identified to resolve this point, thermodynamic argu ments based on observation of equal populations of both dimer types fa vor formation of heterotetramers.