Binding of cystatin C to Alzheimer’s amyloid β inhibits in vitro amyloid fibril formation

The colocalization of cystatin C, an inhibitor of cysteine proteases, with amyloid β (Aβ) in parenchymal and vascular amyloid deposits in brains of Alzheimer’s disease (AD) patients may reflect cystatin C involvement in amyloidogenesis. We therefore sought to determine the association of cystatin C with Aβ. Immunofluorescence analysis of transfected cultured cells demonstrated colocalization of cystatin C and β amyloid precursor protein (βAPP) intracellularly and on the cell surface. Western blot analysis of immunoprecipitated cell lysate or medium proteins revealed binding of cystatin C to full-length βAPP and to secreted βAPP (sβAPP). Deletion mutants of βAPP localized the cystatin C binding site within βAPP to the Aβ region. Cystatin C association with βAPP resulted in increased sβAPP but did not affect levels of secreted Aβ. Analysis of the association of cystatin C and Aβ demonstrated a specific, saturable and high affinity binding between cystatin C and both Aβ1–42 and Aβ1–40. Notably, cystatin C association with Aβ results in a concentration-dependent inhibition of Aβ fibril formation.