title: Identification of the region responsible for fibril formation in the CAD domain of caspase-activated DNase.
abstract: Caspase-activated DNase (CAD) has a compact domain at its N-terminus (CAD domain, 87 amino acid residues), which comprises one alpha-helix and five beta-strands forming a single sheet. The CAD domain of CAD (CAD-CD) forms amyloid fibrils containing alpha-helix at low pH in the presence of salt. To obtain insights into the mechanism of amyloid fibril formation, we identified the peptide region essential for fibril formation of CAD-CD and the region responsible for the salt requirement. We searched for these regions by constructing a series of deletion and point mutants of CAD-CD. Fibril formation by these CAD-CD mutants was examined by fluorescence analysis of thioflavin T and transmission electron microscopy. C-Terminal deletion and point mutation studies revealed that an aromatic residue near the C-terminus (Trp81) is critical for fibril formation. In addition, the main chain conformation of the beta5 strand, which forms a hydrophobic core with Trp81, was found to be important for the fibril formation by CAD-CD. The N-terminal 30 amino acid region containing two beta-strands was not essential for fibril formation. Rather, the N-terminal region was found to be responsible for the requirement of salt for fibril formation.
authors.affiliation: National Institute of Advanced Industrial Science and Technology (AIST), 1-8-31 Midorigaoka, Ikeda, Osaka 563-8577.
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Nakamura
authors.forename: Atsuko
authors.initials: A
authors.lastname: Kobayashi
authors.forename: Yoshihisa
authors.initials: Y
authors.lastname: Hagihara
authors.forename: Tomoyo
authors.initials: T
authors.lastname: Takai
authors.forename: Shouhei
authors.initials: S
authors.lastname: Mine
authors.forename: Toshio
authors.initials: T
authors.lastname: Yamazaki
authors.forename: Noboru
authors.initials: N
authors.lastname: Yumoto
authors.forename: Koichi
authors.initials: K
authors.lastname: Uegaki
mesh_terms: D000682:Amyloid / Q000096:biosynthesis*; D003851:Deoxyribonucleases / Q000737:chemistry* / Q000235:genetics; D006801:Humans; D017354:Point Mutation; D017434:Protein Structure, Tertiary; D012492:Salts; D012965:Sodium Chloride
chemical_list: D000682:Amyloid; D012492:Salts; D012965:Sodium Chloride; D003851:Deoxyribonucleases; C109663:caspase-activated deoxyribonuclease