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Using a monoclonal antibody against the entire C-terminal end of human APP(695) (643-695 sequence) and a monoclonal antibody directed against human beta[1-40] amyloid peptide (betaA), we show the existence of endogenous peptides proteolytically derived from APP in skin exudate of the non transgenic Xenopus laevis frog. The majority of the immunoreactivity is found associated with a 30 kDa molecular species. Biochemical fractionation followed by mass spectrometry identification allowed us to assign this molecular species to C-terminal APP fragments containing all or part of betaA. According to the nature of N- and C-terminal amino acids we identified endogenous beta-, gamma-, epsilon-secretase-like activities, caspase-like activity and numerous endogenous cleavage sites within the beta-amyloid sequence at same sites as those observed in human betaA sequence. All these homologies with human indicate that X. laevis skin exudate is a good natural model to study betaA metabolism. In this way, interestingly, we identified endogenous cleavages at prohormone convertase-like sites not yet described at the same sites in human. Finally, all identified peptide fragments were stably associated with a 20.2 kDa protein. These new observed features suggest new research pathways concerning human betaA metabolism and carriage of hydrophobic peptide fragments issued from APP processing.

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.)