The beta-amyloid peptide, which forms extracellular cerebral deposits in Alzheimer's disease, is derived from a large membrane-spanning glycoprotein referred to as the beta-amyloid precursor protein (APP). The APP is normally cleaved within the beta-amyloid region by a putative proteinase (alpha-secretase) to generate large soluble amino-terminal derivatives of APP, and this event prevents the beta-amyloid peptide formation. It has been suggested that the gelatinase A (matrix metalloproteinase 2, a 72-kDa type IV collagenase) may act either as alpha-secretase or as beta-secretase. Mice devoid of gelatinase A generated by gene targeting develop normally, except for a subtle delay in their growth, thus providing a useful system to examine the role of gelatinase A in the cleavage and secretion of APP in vivo. We show here that APP is cleaved within the beta-amyloid region and secreted into the extracellular milieu of brain and cultured fibroblasts without gelatinase A activity. The data suggest that gelatinase A does not play an essential role in the generation and release of soluble derivatives of APP at physiological conditions.