Analysis of the secondary structure of beta-amyloid (Abeta42) fibrils by systematic proline replacement.
Amyloid fibrils in Alzheimer's disease mainly consist of 40- and 42-mer beta-amyloid peptides (Abeta40 and Abeta42) that exhibit aggregative ability and neurotoxicity. Although the aggregates of Abeta peptides are rich in intermolecular beta-sheet, the precise secondary structure of Abeta in the aggregates remains unclear. To identify the amino acid residues involved in the beta-sheet formation, 34 proline-substituted mutants of Abeta42 were synthesized and their aggregative ability and neurotoxicity on PC12 cells were examined. Prolines are rarely present in beta-sheet, whereas they are easily accommodated in beta-turn as a Pro-X corner. Among the mutants at positions 15-32, only E22P-Abeta42 extensively aggregated with stronger neurotoxicity than wild-type Abeta42, suggesting that the residues at positions 15-21 and 24-32 are involved in the beta-sheet and that the turn at positions 22 and 23 plays a crucial role in the aggregation and neurotoxicity of Abeta42. The C-terminal proline mutants (A42P-, I41P-, and V40P-Abeta42) hardly aggregated with extremely weak cytotoxicity, whereas the C-terminal threonine mutants (A42T- and I41T-Abeta42) aggregated potently with significant cytotoxicity. These results indicate that the hydrophobicity of the C-terminal two residues of Abeta42 is not related to its aggregative ability and neurotoxicity, rather the C-terminal three residues adopt the beta-sheet. These results demonstrate well the large difference in aggregative ability and neurotoxicity between Abeta42 and Abeta40. In contrast, the proline mutants at the N-terminal 13 residues showed potent aggregative ability and neurotoxicity similar to those of wild-type Abeta42. The identification of the beta-sheet region of Abeta42 is a basis for designing new aggregation inhibitors of Abeta peptides.
Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Kyoto 606-8502, Japan.
Akira
A
Morimoto
Kazuhiro
K
Irie
Kazuma
K
Murakami
Yuichi
Y
Masuda
Hajime
H
Ohigashi
Masaya
M
Nagao
Hiroyuki
H
Fukuda
Takahiko
T
Shimizu
Takuji
T
Shirasawa
D000544:Alzheimer Disease / Q000209:etiology / Q000378:metabolism; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D016229:Amyloid beta-Peptides / Q000737:chemistry* / Q000235:genetics; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D066298:In Vitro Techniques; D008854:Microscopy, Electron; D008958:Models, Molecular; D008969:Molecular Sequence Data; D046912:Multiprotein Complexes; D016296:Mutagenesis; D010446:Peptide Fragments / Q000737:chemistry* / Q000235:genetics; D011392:Proline / Q000737:chemistry; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics
D016229:Amyloid beta-Peptides; D046912:Multiprotein Complexes; D010446:Peptide Fragments; D011994:Recombinant Proteins; C075222:amyloid beta-protein (1-42); D011392:Proline