Screening for rabbit brain neuropeptide-metabolizing peptidases. Inhibition of endopeptidase B by bradykinin potentiating peptide 9a (SQ 20881).
Neutral thiol-activated peptidases present in the pH 5-soluble fraction of rabbit brain (separated by step-elution chromatography on diethylaminoethyl cellulose) were screened for the hydrolysis of bradykinin. Lys-bradykinin, Met-Lys-bradykinin, angiotensin I, angiotensin II, substance P, luteinizing hormone-releasing hormone (LH-RH), and neurotensin by bioassay. The column effluent was monitored for bradykinin inactivation and arylamidase activity and combined in six pools on the basis of bradykinin inactivation. The pools were characterized by determining the peptide fragments and amino acids released from bradykinin with an amino acid analyzer. Pools 1 through 3 contained 80% of the kininase activity and essentially all of the endopeptidase A and B activity, whereas pools 4 through 6 accounted for 98% of the recovered arylamidase activity. Bradykinin, angiotensin I, angiotensin II, and substance P were inactivated by all the pools, whereas LH-RH and neurotensin were inactivated by pools 3 and 4, and pools 3, 4, and 5, respectively. These data show that rabbit brain contains peptidases having some selectivity for the inactivation of neuropeptides. Endopeptidase B purified from pool 3 is inhibited by bradykinin-potentiating peptide 9a (BPP9a, SQ 20881) (< Glu-Trp-Pro-Arg-Pro-Gln-Ile-Pro-Pro), a competitive inhibitor of the hydrolysis of bradykinin (Km = 3.5 X 10(-5) M, Ki = 3 X 10(-6) M) which also completely inhibits the inactivation of LH-RH.
D000803:Angiotensin I; D000804:Angiotensin II; D000818:Animals; D001920:Bradykinin; D001921:Brain / Q000201:enzymology*; D007987:Gonadotropin-Releasing Hormone; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000302:isolation & purification*; D007700:Kinetics; D009496:Neurotensin; D009842:Oligopeptides / Q000494:pharmacology*; D011817:Rabbits; D013373:Substance P; D013379:Substrate Specificity; D013722:Teprotide / Q000494:pharmacology*
D007527:Isoenzymes; D009842:Oligopeptides; D000804:Angiotensin II; D013373:Substance P; D007987:Gonadotropin-Releasing Hormone; D009496:Neurotensin; D000803:Angiotensin I; D013722:Teprotide; D001920:Bradykinin