THE CATALYTIC DOMAIN OF INSULIN-DEGRADING ENZYME FORMS A HIGHLY STABLE COMPLEX WITH AMYLOID ƒÒ PEPTIDE

LLOVERA RE; DE TULIO M; ALONSO L; PRAT GAY G; MORELLI L; CASTAÑO EM

Mar del Plata

Congreso; SAIB 43th Annual Meeting; 2007

Insulin-degrading enzyme (IDE) is central to the turnover of insulin and one of the amyloid  (A) proteases in the mammalian brain. Biochemical and genetic data support that IDE may play a role in late onset Alzheimer’s disease (AD).Here we show that a natively folded recombinant IDE was capable of forming a stable complex with A that resisted dissociation after treatment with 70% formic acid, 8M urea or 6M guanidine. This interaction was also observed with rat brain IDE and detected in a SDS-soluble fraction from AD cortical tissue A sequence 17-27, known to be crucial in amyloid assembly was sufficient to form a stable complex with IDE. Monomeric as opposed to aggregated A was competent in complex formation following a very slow kinetics (t1/2 ~45 min). Partial denaturation of IDE as well as pre-incubation with a 10-fold molar excess of insulin prevented complex formation, suggesting that the irreversible interaction of A takes place with at least part of the substrate binding site of the protease. Limited proteolysis showed that A remained bound to a ~23 kDa N-terminal fragment of IDE in an SDS-resistant manner. Mass spectrometry after in gel digestion of the IDE-A17-27complex showed that peptidic fragments derived from the catalytic site of IDE were recovered with A. We propose that the recognition mechanism of IDE towards amyloid forming peptides may lead to a heterologous amyloid-like interaction with novel implications in cerebral amyloidoses.