Angiotensin-1-converting enzyme (ACE) is a zinc metallopeptidase that consists of two homologous catalytic domains (N and C) with different substrate specificities. Angiotesnin I and bradykinin are the most celebrated substrates however, it also hydrolyses the amyloid beta (Aβ) peptide, a causative agent in the progression of Alzheimer's disease. In this study we determined the kinetic parameters of five different forms of human ACE with various Aβ substrates and determined high resolution crystal structures of N-domain in complex with Aβ fragments. For the physiological Aβ(1-16) peptide, a novel ACE cleavage site was found at His14/Gln15. Furthermore, Aβ(1-16) was preferentially cleaved by the truncated N-domain; however, the presence of an inactive C-domain in full-length ACE greatly reduced enzyme activity and affected selectivity. Two fluorogenic substrates, Aβ(4-10)Q and Aβ(4-10)Y underwent endoproteolytic cleavage at the Asp7/Ser8 bond. The Aβ(4-10)Q peptide was a poor substrate of ACE, but was also N-selective. ACE C-domain hydrolysis of Aβ(4-10)Y was effective compared to the other substrates, where hydrolysis under the same conditions did not occur. Despite this improved hydrolysis, Aβ(4-10)Y still displayed N-domain selectivity. Surprisingly, in contrast to Aβ(1-16) and Aβ(4‑10)Q, both the sACE and the double C-domain (CC-sACE) construct showed positive domain cooperativity towards Aβ(4-10)Y. The high resolution crystal structures of the N-domain in complex with five Aβ peptide fragments provided a molecular mechanism of peptide binding and evidence of the enzyme’s broad exoprotease activity. Overall these findings are important for the design of domain selective inhibitors as the differences in domain-selectivity are marked when the truncated domains are compared to the more physiological full length forms.