The hallmark of prion diseases is the cerebral accumulation of a conformationally altered isoform (PrP(Sc)) of a normal cellular protein, the prion protein (PrP(C)). In the inherited form, mutations in the prion protein gene are thought to cause the disease by altering the metabolism of the mutant PrP (PrP(M)) engendering its conversion into PrP(Sc). We used a cell model to study biosynthesis and processing of PrP(M) carrying the glutamic acid to lysine substitution at residue 200 (E200K), which is linked to the most common inherited human prion disease. PrP(M) contained an aberrant glycan at residue 197 and generated an increased quantity of truncated fragments. In addition, PrP(M) showed impaired transport of the unglycosylated isoform to the cell surface. Similar changes were found in the PrP isolated from brains of patients affected by the E200K variant of Creutzfeldt-Jakob disease. Although the cellular PrP(M) displayed some characteristics of PrP(Sc), the PrP(Sc) found in the E200K brains was quantitatively and qualitatively different. We propose that the E200K mutation cause the same metabolic changes of PrP(M) in the cell model and in the brain. However, in the brain, PrP(M) undergoes additional modifications, by an age-dependent mechanism that leads to the formation of PrP(Sc) and the development of the disease.