THE ENDOPEPTIDASE ACTIVITY AND THE ACTIVATION BY CL- OF ANGIOTENSIN-CONVERTING ENZYME IS EVOLUTIONARILY CONSERVED - PURIFICATION AND PROPERTIES OF AN ANGIOTENSIN-CONVERTING ENZYME FROM THE HOUSEFLY, MUSCA-DOMESTICA

A soluble 67 kDa angiotensin-converting enzyme (ACE) has been purified by lisinopril-Sepharose affinity column chromatography from adult hou seflies, Musca domestica. The dipeptidyl carboxypeptidase activity tow ards benzoyl-Gly-His-Leu was inhibited by captopril (IC50 50 nM) and f osinoprilat (IC50 251 nM), two inhibitors of mammalian ACE, and was ac tivated by Cl- (optimal Cl- concentration 600 mM). Musca ACE removed C -terminal dipeptides from angiotensin I, bradykinin, [Leu(5)]enkephali n and [Met(5)]enkephalin and also functioned as an endopeptidase by hy drolysing dipeptideamides from [Leu(5)]enkephalinamide and [Met(5)]enk ephalinamide, and a dipeptideamide and a tripeptideamide from substanc e P. Musca ACE was also able to cleave a tripeptide from both the N-te rminus and C-terminus of luteinizing hormone-releasing hormone, with C -terminal hydrolysis predominating. Maximal N-terminal tripeptidase ac tivity occurred at 150 mM NaCl, whereas the C-terminal tripeptidase ac tivity continued to rise with increasing concentration of Cl- (0-0.5 M ). Musca ACE displays properties of both the N- and C-domains of human ACE, indicating a high degree of conservation during evolution of the substrate specificity of ACE and its response to Cl-.