Limited proteolysis of human kidney angiotensin-converting enzyme and generation of catalytically active N- and C-terminal domains.

The somatic form of angiotensin converting enzyme is a class I ectoenzyme that is bound to the surface of endothelial calls. It consists of two homologous, catalytic domains of approximately 600 residues each; a juxtamembrane "stalk" region; a transmembrane, hydrophobic sequence; and a 30 residue, C-terminal cytosolic domain. We have used limited proteolysis to probe the structural and functional properties of the enzyme. Endoproteinase Asp-N cleaves both the Thr615-Asp616 and the Leu1219-Asp1220 peptide bonds to generate the two catalytic domains which were isolated by a combination of immunoaffinity and lisinopril Sepharose affinity chromatography. The enzymatic characteristics of the N and C fragments were examined with angiotensin I, hippuryl-His-Leu, and luteinizing hormone-releasing hormone and indicate that both fragments contain catalytically active sites that retain their individual functional integrity.