Substance P and [Leu]enkephalin are hydrolyzed by an enzyme in pig caudate synaptic membranes that is identical with the endopeptidase of kidney microvilli.

Abstract

The hydrolysis of [Leu]enkephalin and substance P by purified pig kidney endopeptidase (EC 3.4.24.11) and synaptic membranes prepared from pig caudate nuclei has been compared. The hydrolysis of an enkephalin analogue (Tyr-D-Ala-GlyPhe-Leu) at the Gly-Phe bond was completely inhibited by phosphoramidon. The IC50 concentration (8 nM) was similar to that reported for [Leu] Biochem. J. 203,[519][520][521][522]. Seven peptides were produced when substance P (Arg-Pro-Lys-Pro-Gln-Gln-Phe-Phe-GlyLeu-Met-NH2) was hydrolyzed by the kidney endopeptidase. These were formed by cleavage at bonds Gln-Phe (positions 6 and 7), Phe-Phe (positions 7 and 8), and Gly-Leu (positions 9 and 10). Synaptic membranes generated peptides with the same HPLC retention times and hydrolysis of substance P by either preparation was inhibited completely by 10 FM phosphoramidon. The most susceptible bond appeared to be Gly-Leu (positions 9 and 10). A specific polyclonal antibody raised in rabbits to purified pig endopeptidase inhibited the hydrolysis of [Leu]enkephalin and substance P by detergent-solubilized kidney microvilli or synaptic membranes; the titration curves were essentially identical. We conclude that the endopeptidase, which we suggest should be designated "endopeptidase-24.11," is present in caudate synaptic membranes and could play an important role in the hydrolysis of neuropeptides.When the inactivation of biologically active peptides has been reported it has become commonplace to name the peptidase responsible in a way that implies the existence of peptide-specific enzymes. Although peptidases of high specificity do exist, such a close relationship between peptidase and substrate is unusual. The field of neuropeptide metabolism in brain has, in particular, been plagued by this restrictive nomenclature [e.g., enkephalinase and substance P-degrading enzyme (1-4)]-a limitation that has consequently hindered comparisons with wellcharacterized peptidases from other tissues. It is our contention (5) that a limited number of peptidases mediate a wide range of functions and that their cellular and subcellular localization, rather than peptide specificity, defines their roles at different sites.

The Enhancing Effects of Amastatin, Phosphoramidon and Captopril on the Potency of [Met5]-Enkephal n in Rat Vas Deferens

“…The IC50 value of enkephalin in the presence of 1 /-,M of phosphoramiclon was not significantly different from that of 2 uM of phosphoramidon, but significantly lower than that of 0.1 /iM of phosphoramidon (Table 3). This indicated that the enkephalin hydrolyzing endopeptidase-24.11 in rat vas deferens was inhibited maximally with phos phoramidon at the final concentration of 1 riM, being consistent with the previous results obtained with guinea-pig ileum (1 ), mouse vas deferens (2), pig caudate synaptic membranes (12) and purified endopeptidase 24.11 from pig kidney (13).…”

The Role of Bestatin-Sensitive Aminopeptidase, Angiotensin Converting Enzyme and Thiorphan-Sensitive "Enkephalinase" in the Potency of Enkephalins in the Guinea-Pig Ileum

“…peptidase inhibitors employed (dotted arrows). "Enkephalinase" has recently been reported to be identical with endopeptidase 24.11 (10).…”

Canine bombesin‐like gastrin releasing peptides stimulate gastrin release and acid secretion in the dog.

“…11 efficiently inactivates gastrin releasing peptide 10 by cleavage of the His8-Leu9 bond (Bunnett, Kobayashi, Orloff, Turner, Reeve & Walsh, 1985). This membrane-bound peptidase is present in many tissues but is concentrated particularly in the kidney microvilli (Matsas, Fulcher, Kenny & Turner, 1983).…”