RESUMEN
UNLABELLED: The aggregates of amyloid beta peptides (Aßs) are regarded as one of the main pathological hallmarks of Alzheimer's disease (AD). An imbalance between the rates of synthesis and clearance of Aßs is considered to be a possible cause for the onset of AD. Dipeptidyl peptidases II and IV (DPPII and DPPIV) are serine proteases removing N-terminal dipeptides from polypeptides and proteins with proline or alanine on the penultimate position. Alanine is an N-terminal penultimate residue in Ðßs, and we presumed that DPPII and DPPIV could cleave them. The results of present in vitro research demonstrate for the first time the ability of DPPIV to truncate the commercial Aß40 and Aß42 peptides, to hinder the fibril formation by them and to participate in the disaggregation of preformed fibrils of these peptides. The increase of absorbance at 334 nm due to complex formation between primary amines with o-phtalaldehyde was used to show cleaving of Aß40 and Aß42. The time-dependent increase of the quantity of primary amines during incubation of peptides in the presence of DPPIV suggested their truncation by DPPIV, but not by DPPII. The parameters of the enzymatic breakdown by DPPIV were determined for Aß40 (Km=37.5 µM, kcat/Km=1.7×10(3)M(-1)sec(-1)) and Aß42 (Km=138.4 µM, kcat/Km=1.90×10(2)M(-1)sec(-1)). The aggregation-disaggregation of peptides was controlled by visualization on transmission electron microscope and by Thioflavin-T fluorescence on spectrofluorimeter and fluorescent microscope. DPPIV hindered the peptide aggregation/fibrillation during 3-4 days incubation in 20mM phosphate buffer, pH 7.4, 37°C by 50-80%. Ovalbumin, BSA and DPPII did not show this effect. In the presence of DPPIV, the preformed fibrils were disaggregated by 30-40%. CONCLUSION: for the first time it was shown that the Aß40 and Aß42 are substrates of DPPIV. DPPIV prohibits the fibrillation of peptides and promotes disaggregation of their preformed aggregates.
