IDENTIFICATION AND PROPERTIES OF A PEPTIDYL DIPEPTIDASE IN THE HOUSEFLY, MUSCA-DOMESTICA, THAT RESEMBLES MAMMALIAN ANGIOTENSIN-CONVERTING ENZYME

[D-Ala(2),Leu(5)]Enkephalin was readily metabolized by membranes (40000 g pellet) prepared from heads of the housefly, Musca domestica, with Gly(3)-Phe(4) being the major site of cleavage. This hydrolysis was only partially inhibited (40%) by 10 mu M phosphoramidon, an inhibitor of endopeptidase-24.11, but was almost totally abolished in the presence of a mixture of 10 mu M phosphoramidon and 10 mu M captopril, a potent inhibitor of mammalian angiotensin-converting enzyme (ACE). An assay for ACE employing Bz-Gly-His-Leu as the substrate was used to confirm the presence of an ACE-like peptidyl dipeptidase activity in fly head membranes. The peptidase had a K-m of 1.91 mM for Bz-Gly-His-Leu and a pH optimum of 8.2. The activity was inhibited by 100 mu M EDTA and was greatly activated by ZnCl2 but not other bivalent metal ions. Captopril, lisinopril, fosinoprilat and enalaprilat, all selective inhibitors of mammalian ACE, were also good inhibitors of the insect enzyme with IC50 values of 400 nM, 130 nM, 16 nM and 290 nM respectively. An M(r) value of around 87000 was obtained for this enzyme from gel-filtration chromatography, indicating that the insect enzyme is similar in size to mammalian testicular ACE (M(r) = 9000- 110000) and not the larger form of the enzyme (M(r) = 150000-180000) found in mammalian somatic tissues. The fly peptidyl dipeptidase was released from membranes into a soluble fraction by incubating the head membranes at 37 degrees C but not at 0 degrees C, suggesting that the insect ACE-like enzyme can be solubilized from cell surfaces through the activity of a membrane-bound enzyme activity. In conclusion, we have shown the existence of a peptidyl dipeptidase in membranes from the heads of M. domestica, which has similar properties to those of mammalian ACE.