Water-Soluble β-Sheet Models Which Self-Assemble into Fibrillar Structures

Abstract

Self-assembly of beta-sheet domains resulting in the formation of pathogenic, fibrillar protein aggregates (amyloids) is a characteristic feature of various medical disorders. These include neurodegenerative diseases, such as Alzheimer's, Huntington's, and Creutzfeldt-Jacob's. A significant problem in studying such aggregation processes is the poor solubility of these beta-sheet complexes. The present work describes water-soluble de novo beta-sheet peptides which self-assemble into fibrillar structures. The model peptides enable studies of the relationship between beta-sheet stability and association behavior. The peptides [DPKGDPKG-(VT)n-GKGDPKPD-NH2, n = 3-8] are composed of a central beta-sheet-forming domain (VT-sequence), and N- and C-terminal nonstructured octapeptide sequences which promote water solubility. Conformational analyses by circular dichroism and Fourier transform infrared spectroscopy indicate the influence of peptide length, D-amino acid substitution, and concentration on the ability of the peptides to form stable beta-sheet structures. The association behavior investigated by analytical ultracentrifugation and dynamic light scattering was found to correlate strongly with the stability of a beta-sheet conformation. Model peptides with n >/= 6 form stable, water-soluble beta-sheet complexes with molecular masses of more than 2000 kDa, which are organized in fibrillar structures. The fibrils examined by Congo Red staining and electron microscopy show some similarities with naturally occurring amyloid fibrils.

Time-Course and Regional Analyses of the Physiopathological Changes Induced after Cerebral Injection of an Amyloid β Fragment in Rats

“…4 Structure-activity studies revealed that peptides containing the highly hydrophobic 25-35 region formed stable aggregates and mediated neuronal death by necrosis or apoptosis. 5,6,7 The truncated A␤ [25][26][27][28][29][30][31][32][33][34][35] fragment includes extracellular and intramembrane residues that have been reported to represent an active region of A␤. 8 In vivo, two nontransgenic rodent models of AD have been studied to analyze the molecular, morphological, and behavioral consequences of amyloid toxicity: the infusion of A␤ 1-40/42 protein and the injection of preaggregated A␤ [25][26][27][28][29][30][31][32][33][34][35] peptide.…”

“…These models have led to highly pertinent pathomimetic observations and provide complementary approaches to the numerous transgenic mouse lines developed as AD models. In particular, at 1 week after A␤ [25][26][27][28][29][30][31][32][33][34][35] injection, reactive gliosis was observed in the rat hippocampus, 9 caspase-3 activity was induced in the hippocampus and cortex, 9 a reduction in the number of neurons was measured in the CA1 or CA3 hippocampal area, 9 -11 and significant oxidative stress was observed. [11][12][13] Nonetheless, this amyloid toxicity model remains controversial with respect to AD.…”

“…First, although the presence of A␤ [25][26][27][28][29][30][31][32][33][34][35] has been repeatedly demonstrated in brains of AD patients, 14,15 this fragment appears to be a minor peptide in plaques and its contribution to the toxicity induced by all amyloid-related species remains unclear. Second, the intracerebroventricular injection of A␤ [25][26][27][28][29][30][31][32][33][34][35] induces marked toxicity within days or weeks, which is not in accord with the long-term progression of AD in humans (an argument that is unrelated to the nature of the toxicity). Third, it is possible that peptide injection mimics only the amyloid toxicity observed in AD physiopathology, but not the processes involved in A␤ formation or Tau protein hyperphosphorylation.…”

“…Klementiev et al 16 recently documented some evidence that contradicts this latter reservation. Moreover, very recently, Chavant et al 17 reported that, at 2 weeks after injection, A␤ [25][26][27][28][29][30][31][32][33][34][35] significantly increased APP processing in the hippocampus of mice.…”

“…We therefore analyzed the consequences of intracerebroventricular injection of aggregated A␤ [25][26][27][28][29][30][31][32][33][34][35] in terms of time-course changes and in different brain structures through several morphological, hormonal, biochemical, and behavioral parameters, as well as addressing the modifications in central APP processing, brain-derived neurotrophic factor (BDNF) levels, cerebral inflammation, and hippocampus vulnerability. We also evaluated over time the distribution within brain structures of the injected A␤ [25][26][27][28][29][30][31][32][33][34][35] fragment.…”