The betabellin structure is a de novo designed beta-sandwich protein consisting of two 32-residue beta-sheets packed against one another by hydrophobic interactions. d-Amino acid residues are used to energetically favor formation of type-I' beta turns. Air oxidation of betabellin 15S (B15S) (HSLTAKIpkLTFSIAphTYTCAVpkYTAKVSH, where p denotes d-Pro, h denotes d-His, and k denotes d-Lys) yields betabellin 15D (B15D), a 64-residue disulfide-bridged protein. The amino acid sequence of B15D contains a conformationally constrained d-Pro residue at the i + 1 position of each type-I' beta turn. To test whether d-Pro residues are necessary for folding at these positions, the six d-Pro residues of B15D are replaced by d-Ala residues in betabellin 16D (B16D). Previously, transmission electron microscopy showed that B15D forms unbranched, 35-A wide fibrils that associate into bundles in 5.0 mM 3-(N-morpholino)propanesulfonate and 250 mM NaCl at pH 7; under these conditions, B16D forms ribbon-like assemblies. The B15D fibrils resemble the protofilaments that constitute amyloid fibrils. The present studies show that both B15D and B16D have characteristics of amyloidogenic proteins: the unbranched fibrils and ribbons stained with Congo red and displayed a green birefringence, exhibited a cross-beta structure, and bound 1-anilino-8-naphthalenesulfonate. Thus, these de novo designed beta-sandwich proteins should provide useful models for studying the mechanism of amyloid protofilament formation and assembly into amyloid fibrils and for designing potential inhibitors of amyloidogenesis.
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Lim et al. (2000) studied this question.