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The presence of senile plaques in the brain is one of the pathological hallmarks of Alzheimer's disease (AD). The biogenesis and clearance of the amyloid β peptide (A β ), the main component of the lesions, lie at the center of the pathogenesis of AD. In sporadic AD, the increase of A β levels seems to be indicative of failure of clearance mechanisms. We previously showed that the clearance of the wild type A β40 peptide by various neuronal and non-neuronal cells occurs through a same proteolytic process and that A β degradation was primarily dictated by its conformational state (Panchal et al., 2007). To gain further insights on the role of the peptide conformation in the clearance mechanism of A β , two A β40 peptides, known to be associated with amyloid angiopathy (Dutch and Flemish mutations), and the rodent A β40 peptide were catabolized by several cells by using the same experimental approach. The peptide fragments, generated by proteolytic cleavage of substrates in cell supernatants, were identified by LC-MS and the cleavage sites of proteases were deduced. In parallel, conformational states of wild type A β 40 peptide and of the three A β 40 variants were characterized by circular dichroism spectroscopy. We provide data suggesting that discrete conformational changes of A β 40 peptide regulate its clearance rate by neuronal and non-neuronal cells.

Deposition of amyloid-beta peptide (Abeta) in the brain is an early and invariant feature of all forms of Alzheimer's disease (AD). As for all proteins or peptides, the steady-state level of Abeta peptide is determined not only by its production, but also by its degradation. So, overactive proteases involved in generating Abeta from amyloid precursor protein or underactive Abeta-degrading enzymes could lead to abnormal Abeta deposition in the brain. Since in the sporadic forms of AD (90% of all AD cases) an impaired clearance of Abeta appears to be at the origin of its aggregation and tissue deposition, we have investigated its proteolytic degradation by several neuronal and non-neuronal cells. In this report, we show that these cell types exhibit a similar profile of Abeta-degradation by cell-surface and secreted proteases which were respectively characterized as metallo- and serine proteases. By using a combination of the liquid chromatography/on-line mass spectrometry, we demonstrate that: (i)-the membrane associated protease(s) hydrolizes Abeta40 essentially at Lys(28) Gly(29), Phe(19) Phe(20) and Val(18) Phe(19) bonds; and (ii)-the secreted protease(s) cleaves the generating fragments Abeta (1-28), Abeta (1-19), Abeta (1-18) at His(14) Gln(15) bond and also Abeta (1-28) at Phe(20) Ala(21) and Asp(23) Val(24) sites. This is the first time our results define a proteolytic degradation process of Abeta40 that appears to be independent of the cell type and may represent a general pattern of its enzymatic clearance.

Dactylysin (EC 3.5.24.60) is a metalloendopeptidase first isolated from the skin granular gland secretions of Xenopus laevis. This peptidase hydrolyzes bonds on the amino-terminus of singlets and between doublets of hydrophobic amino acids and was considered to play a role in the in vivo inactivation of biologically active regulatory peptides. Here, we show that dactylysin has also the ability to cleave human beta[1-40]-amyloid peptide and related peptides. Cleavage of the wild type beta[1-40]-amyloid peptide form, and to a lesser extent Flemish and Dutch mutants, occurred predominantly at the His14-Glu15 bond. We demonstrate that frog skin exudate contains a full-length amyloid protein precursor detected by immunochemical cross-reactivity with monoclonal antibody against C-terminal human amyloid protein precursor. The possibility that dactylysin, might be involved in normal catabolism of beta amyloid peptide of Xenopus laevis is discussed., (Copyright 2002 Elsevier Science Inc.)

Many chemokines have direct suppressive activity in vitro and in vivo on primitive hematopoietic cells. However, few chemokine-derived peptides have shown a significant activity in inhibiting hematopoiesis. Interestingly, a peptide derived from the 34-58 sequence of the CXC chemokine platelet factor 4 (PF4) produced a 30-40% inhibition of proliferation of murine hematopoietic progenitors (CFU-MK, CFU-GM, and BFU-E) in vitro, at concentrations of 30-60-fold lower than PF4. The aim of the present work was to define the structural parameters and motifs involved in conferring biological activity to the peptide PF4(34-58). Both structural predictions and determinations revealed a new helical motif that was further localized between residues 38 and 46. This helix was necessary for binding of the peptide and for permitting the functional DLQ motif at position 54-56 to activate the putative receptor site. Peptides lacking either the helical or the DLQ motif were devoid of inhibitory activity on the hematopoietic progenitors in vitro. However, among inactive peptides, only those having the helical motif counteracted the inhibition induced by the active peptide PF4(34-58). This suggested that the helix might be required for peptide interactions with a putative receptor site, whereas the DLQ motif would be implicated in the activation of this receptor. These results identify for the first time the dual requirements for the design of chemokine-derived peptides with high suppressive activity on hematopoiesis, as well as for the design of molecules with antagonistic action.