Enzyme kinetic studies and inhibition by oligopeptides of LH-RH degradation in rat hypothalamus and pituitary.
The enzyme kinetic parameters of the degradation of luteinizing hormone-releasing hormone (LH-RH) and L-cystine-bis-(4-nitroanilide) (Cys-NA) by rat hypothalamic (HYP) and pituitary (PIT) extracts and the effect of various oligopeptides on the rate of LH-RH inactivation were investigated in vitro. The 105,000 x g supernatant of 1 rat HYP inactivated 57 microgram LH-RH during a 30 min incubation (Km = 12.4 microM, V max = 2.33 microgram LH-RH/mg protein/min), and of one rat anterior PIT, 48 microgram LH-RH during 30 min of incubation (Km = 12.2 microM, V max = 8.0 microgram LH-RH/mg protein/min). The synthetic substrate Cys-NA competitively inhibited LH-RH degradation with a Ki of 8.5 microM in the HYP and 6 microM in the PIT enzyme preparation. Vice versa, LH-RH also competitively inhibited the cleavage of Cys-NA with inhibition constants of 14 microM (HYP) and 15 microM (PIT) indicating that the 2 substrates are probably cleaved by the same enzyme. The most effective inhibitors of LH-RH degradation were found to be angiotensin-related peptides, neurotensin, bradykinin, and bacitracin. A relatively weak effect was obtained with oxytocin, enkephalin and puromycin. It is concluded that endogenous oligopeptides such as angiotensins, neurotensin, bradykinin, etc., may possibly influence H-RH degradation in the PIT and the HYP. The synthetic substrate Cys-NA may be an appropriate substrate for measuring the activity of an LH-RH-degrading peptidase, which therefore could be classified as arylamidase.
D000626:Aminopeptidases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D005260:Female; D007987:Gonadotropin-Releasing Hormone / Q000378:metabolism*; D007031:Hypothalamus / Q000201:enzymology / Q000378:metabolism*; D007700:Kinetics; D009842:Oligopeptides / Q000494:pharmacology*; D010902:Pituitary Gland / Q000201:enzymology / Q000378:metabolism*; D051381:Rats; D013379:Substrate Specificity
D000900:Anti-Bacterial Agents; D009842:Oligopeptides; D007987:Gonadotropin-Releasing Hormone; D000626:Aminopeptidases