Parkinson's disease is associated with the aberrant aggregation of alpha-synuclein. Although the causes of this process are still unclear, post-translational modifications of alpha-synuclein are likely to play a modulatory role. Since alpha-synuclein is constitutively N-terminally acetylated, we investigated how this post-translational modification alters the aggregation behavior of this protein. By applying a three-pronged aggregation kinetics approach, we observed that N-terminal acetylation results in a reduced rate of lipid-induced aggregation and slows down both elongation and fibrilcatalyzed aggregate proliferation. An analysis of the amyloid fibrils produced by the aggregation process revealed different morphologies for the acetylated and nonacetylated forms in both lipid-induced aggregation and seed-induced aggregation assays. In addition, we found that fibrils formed by acetylated alpha-synuclein exhibit a lower beta-sheet content. These findings indicate that N-terminal acetylation of alpha-synuclein alters its lipid-dependent aggregation behavior, reduces its rate of in vitro aggregation, and affects the structural properties of its fibrillar aggregates.