Betabellins 15D and 16D, de Novo designed beta-sandwich proteins that have amyloidogenic properties.
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.
Department of Chemistry, The University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, 27599, USA.
A
A
Lim
A M
AM
Makhov
J
J
Bond
H
H
Inouye
L H
LH
Connors
J D
JD
Griffith
B W
BW
Erickson
D A
DA
Kirschner
C E
CE
Costello
D000595:Amino Acid Sequence; D000682:Amyloid; D000686:Amyloidosis; D000817:Anilino Naphthalenesulfonates; D004396:Coloring Agents; D003224:Congo Red; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000737:chemistry* / Q000378:metabolism; D011487:Protein Conformation; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D011994:Recombinant Proteins; D013050:Spectrometry, Fluorescence; D014961:X-Ray Diffraction
D000682:Amyloid; D000817:Anilino Naphthalenesulfonates; D004396:Coloring Agents; D009842:Oligopeptides; D011506:Proteins; D011994:Recombinant Proteins; C114709:betabellin 15; C400381:betabellin 16D; D003224:Congo Red