Abstract

Many studies suggest that BACE 1 is the genuine β-secretase; however, this is not undisputed. The wild-type (WT) β-site of the amyloid precursor protein (APP) present in the worldwide population is cleaved very slowly (k _cat/K _m: approx. 50 m ^-1 s^-1), while proteases acting on relevant substrates are much more efficient (k _cat/K _m: 10⁴–10⁶ m ^-1 s^-1). Knock-out of BACE 1 in mouse markedly reduces Aβ formation. Nevertheless, studies in other systems show that knock-out experiments in rodents and corresponding genetic defects in human may reveal different phenotypes. Considering these issues, we searched for other β-secretase candidate(s), identified cathepsin D, and evaluated properties of cathepsin D related to BACE 1 that were not examined previously. The kinetic constants (k _cat, K _m, k _cat/K _m) for cleaving peptides with β-sites of the WT or the mutated Swedish families (SW) APP by human BACE 1 and cathepsin D were determined and found to be similar. Western blots reveal that in human brain cathepsin D is approximately 280-fold more abundant than BACE 1. Furthermore, pepstatin A strongly inhibits the cleavage of SW and WT peptides by both brain extracts and cathepsin D, but not by BACE 1. These findings indicate that β-secretase activity observed in brain extracts is mainly due to cathepsin D. Nevertheless, as both BACE 1 and cathepsin D show poor activity towards the WT β-site sequence, it is necessary to continue the search for additional β-secretase candidate(s).