Degradation of alpha and beta neo-endorphin by rat brain membrane peptidases

Fractionation of Triton-solubilized rat brain membranes on diethylaminoethyl-cellulose resolves two peptidases which hydrolyze beta-neo-endorphin. One of these peptidases was identified as Angiotensin Converting Enzyme by (a) its sensitivity to inhibition by the specific inhibitors MK422 and captopril, (b) by the identification of reaction products, and (c) by comparison to authentic angiotensin converting enzyme. In contrast, alpha-neo-endorphin hydrolysis by angiotensin converting enzyme could not be detected. The second enzyme active on beta-neo-endorphin was identified as an aminopeptidase. This aminopeptidase is identical to the previously described enkephalin-degrading aminopeptidase. The possible involvement of these enzymes in the metabolism of opioid peptides is discussed.     

Angiotensin converting enzyme has an inhibitory role in CGRP metabolism in human skin

The neutral endopeptidase (NEP) is important for calcitonin gene related peptide (CGRP) degradation, while the role of angiotensin converting enzyme (ACE) remains unclear. By using dermal microdialysis we explored the effect of phosphoramidon (NEP blocker), captopril (ACE blocker) and a mixture of both drugs on the intensity of electrically-induced CGRP-mediated neurogenic flare. The results reveal that phosphoramidon elevated flare intensity, but that this was not further increased by adding captopril. In contrast, neurogenic flare was decreased when the drug mixture was applied in compared to NEP only. Electrically released CGRP levels could be measured directly in perfusates containing phosphoramidon and the mixture. Again, CGRP levels were elevated in phosphoramidon treated sites, and significantly reduced upon adding captopril. These findings suggest that NEP and ACE do not have additive effects regarding neuropeptide degradation. In contrast, inhibition of ACE seems to augment CGRP catabolism.     

Purification and Characterization of a Peptidyl Dipeptidase Resembling Angiotensin Converting Enzyme from the Electric Organ of Torpedo marmorata

The electric organ of Torpedo marmorata contains a membrane-bound, captopril-sensitive metallopeptidase that resembles mammalian angiotensin converting enzyme (peptidyl dipeptidase A; EC 3.4.15.1). The Torpedo enzyme has now been purified to apparent homogeneity from electric organ by a procedure involving affinity chromatography using the selective inhibitor lisinopril immobilised to Sepharose via a 28-A spacer arm. The purified protein, like the mammalian enzyme, acted as a peptidyl dipeptidase in cleaving dipeptides from the C-terminus of a variety of peptide substrates, including angiotensin I, bradykinin, [Met5]enkephalin, [Leu5]enkephalin, and the model substrate hippuryl (benzoylglycyl; BzGly)-His-Leu. The hydrolysis of BzGly-His-Leu was activated by Cl-. Enzyme activity was inhibited by classical angiotensin converting enzyme inhibitors, including captopril, enalaprilat (MK422), and lisinopril (MK521). Torpedo angiotensin converting enzyme, like its mammalian counterpart, was also able to act as an endopeptidase in hydrolysing the amidated neuropeptide substance P. Hydrolysis of substance P occurred primarily at the Phe8-Gly9 bond with release of the C-terminal tripeptide, Gly-Leu-MetNH2, and this hydrolysis was blocked by selective inhibitors. The Torpedo enzyme was recognised by a polyclonal antibody to pig kidney angiotensin converting enzyme on immunoelectrophoretic (Western) blot analysis. Thus, on the basis of substrate specificity, inhibitor sensitivity, and immunological criteria, the Torpedo enzyme closely resembles mammalian angiotensin converting enzyme. However, the Torpedo enzyme appears somewhat larger (Mr = 190,000) than the pig kidney enzyme (Mr = 180,000) on sodium dodecyl sulphate-polyacrylamide gel electrophoresis. The endogenous peptide substrate(s) for Torpedo electric organ angiotensin converting enzyme and the physiological role of the enzyme in this tissue remain to be evaluated.     

Metalloendopeptidase (EC 3.4.24.11) but not angiotensin converting enzyme is involved in the inactivation of substance P by synaptic membranes of the rat substantia nigra

Substance P is a neuropeptide released in vivo from the substantia nigra, the principal substance P nerve terminal region in the rat brain. Its inactivation was investigated in a purified nigral synaptic membrane preparation. The membrane-bound enzyme shares many features with the endopeptidase 24-11 (EC 3.4.24.11): 1) hydrolysis of peptide bonds Gln6-Phe7, Phe7-Phe8 and Gly9-Leu10, 2) sensitivity to the inhibition by phosphoramidon and 3) relative affinity for substance P. Bestatine and captopril inhibit only the hydrolysis of the metabolites. These results suggest that substance P is inactivated in substantia nigra by endopeptidase 24-11 and that a bestatin-sensitive aminopeptidase and angiotensin converting enzyme may play a role in subsequent degradation of the substance P metabolites.     

Catabolism of gastrin releasing peptide and substance P by gastric membrane-bound peptidases

The catabolism of two gastric neuropeptides, the C-terminal decapeptide of gastrin releasing peptide-27 (GRP10) and substance P (SP), by membrane-bound peptidases of the porcine gastric corpus and by porcine endopeptidase-24.11 ("enkephalinase") has been investigated. GRP10 was catabolized by gastric muscle peptidases (specific activity 1.8 nmol min-1 mg-1 protein) by hydrolysis of the His8-Leu9 bond and catabolism was inhibited by phosphoramidon (I50 approx. 10(-8) M), a specific inhibitor of endopeptidase-24.11. The same bond in GRP10 was cleaved by purified endopeptidase-24.11, and hydrolysis was equally sensitive to inhibition by phosphoramidon. SP was catabolized by gastric muscle peptidases (specific activity 1.7 nmol min-1 mg-1 protein) by hydrolysis of the Gln6-Phe7, Phe7-Phe8 and Gly9-Leu10 bonds, which is identical to the cleavage of SP by purified endopeptidase-24.11. The C-terminal cleavage of GRP10 and SP would inactivate the peptides. It is concluded that a membrane-bound peptidase in the stomach wall catabolizes and inactivates GRP10 and SP and that, in its specificity and sensitivity to phosphoramidon, this peptidase resembles endopeptidase-24.11.     

3H]Captopril binding to membrane associated angiotensin converting enzyme

[3H]Captopril binding to membrane fractions of rat tissues is saturable and reversible with a KD of 2.4 nM. [3H]Captopril binding and angiotensin converting enzyme measured with hippuryl-L-histidine-L-leucine are distributed in parallel between different tissues and brain regions, with highest levels in the choroid plexus, lung and corpus striatum. Captopril, N-(1(S)-carboxy-3-phenyl-propyl)-L-alanyl-L-proline, N-(1(S)-carboxy-3-phenyl-propyl)-L-lysyl-L-proline, teprotide, thiorphan and S-acetylcaptopril each have similar potencies for inhibition of [3H]captopril binding and of angiotensin converting enzyme. These data strongly indicate that [3H]captopril binds selectively to angiotensin converting enzyme. [3H]Captopril binding evaluation should help clarify the localization and function of angiotensin converting enzyme and assist in defining pharmacologic actions of captopril.     

Catabolism of substance P in the stomach wall of the rat

The aim of this study was to examine the catabolism of substance P (SP) in the stomach wall of the rat. Catabolism in vitro was investigated by incubation of unlabelled and tritiated SP (prolyl 2,4-3,4(n)-3H SP) with membrane bound-peptidases prepared from the rat gastric corpus. Catabolism was studied in vivo by use of a catheter chronically implanted in the stomach wall to deliver tritiated SP to the gastric tissues and implanted dialysis fibers to collect the catabolic products. The products from both experiments were separated by high pressure liquid chromatography and identified by their retention times or amino acid analysis. Membrane-bound peptidases in vitro hydrolyzed both unlabelled and tritiated SP and the products of hydrolysis were consistent with the cleavage of three bonds: Gln6-Phe7, Phe7-Phe8 and Gly9-Leu10. None of the peptide fragments would be expected to be biologically active. Only those fragments with tritiated Pro residues could be detected in vivo. The major identified products were SP(1-2) and SP(3-4), with smaller amounts of SP(1-4), SP(1-6), SP(1-7), SP(1-8) and SP(1-9). The enzymes that may be responsible for these cleavage patterns are discussed.     

Peptidases involved in the catabolism of neurotensin: inhibitor studies using superfused rat hypothalamic slices

In order to identify which peptidases are involved in the catabolism of neurotensin in the CNS, [3H-Tyr3,11]-neurotensin was superfused over rat hypothalamic slices in the presence and absence of peptidase inhibitors. The degree of degradation of the peptide was determined by reverse phase HPLC separation of 3H-labelled neurotensin from 3H-labelled products. Very little degrading activity was released from the slice into the medium during the superfusion. In the absence of inhibitors, 20 to 50% of 3H-neurotensin was degraded giving mainly 3H-Tyr along with other unidentified 3H-labelled products. Inhibitors of endopeptidase 24.11 (phosphoramidon) and proline endopeptidase (antibody) had no effect on the degradation. Captopril, an inhibitor of angiotensin converting enzyme, had a small inhibitory effect. In contrast, dynorphin(1-13), an inhibitor of a soluble, thiol dependent metallopeptidase which hydrolyses neurotensin at Arg8-Arg9, gave greater than 80% inhibition of 3H-neurotensin degradation in the slice preparation. 1,10-Phenanthroline, an inhibitor of metallopeptidases, was also an effective inhibitor. The dynorphin sequence responsible for the inhibition contains the Arg6-Arg7 bond. Other peptides (bradykinin and angiotensin) which are substrates of the soluble metallopeptidase also inhibited neurotensin breakdown by the slice. This evidence suggests that this thiol dependent metalloendopeptidase is the major neurotensin catabolizing enzyme in hypothalamic slices.     

Inactivation of Neurotensin by Rat Brain Synaptic Membranes. Cleavage at the Pro10-Tyr11 Bond by Endopeptidase 24.11 (Enkephalinase) and a Peptidase Different from Proline-Endopeptidase

It was shown previously that the tridecapeptide neurotensin is inactivated by rat brain synaptic membranes and that one of the primary inactivating cleavages occurs at the Pro10-Try11 peptide bond, leading to the formation of NT1-10 and NT11-13. The present study was designed to investigate the possibility that this cleavage was catalyzed by proline endopeptidase and/or endopeptidase 24.11 (enkephalinase). Purified rat brain synaptic membranes were found to contain a N-benzyloxycarbonyl-Gly-Pro-4-methyl-coumarinyl-7-amide-hydrolyzin g activity that was markedly inhibited (93%) by the proline endopeptidase inhibitor N-benzyloxycarbonyl-Pro-Prolinal and partially blocked (25%) by an antiproline endopeptidase antiserum. In contrast, the cleavage of neurotensin at the Pro10-Tyr11 bond by synaptic membranes was not affected by N-benzyloxycarbonyl-Pro-Prolinal and the antiserum. When the conversion of NT1-10 to NT1-8 by angiotensin converting enzyme was blocked by captopril and when the processing of NT11-13 by aminopeptidase(s) was inhibited by bestatin, it was found that thiorphan, a potent endopeptidase 24.11 inhibitor, partially decreased the formation of NT1-10 and NT11-13 by synaptic membranes. In conclusion: (1) proline endopeptidase, although it is present in synaptic membranes, is not involved in the cleavage of neurotensin at the Pro10-Tyr11 bond; (2) endopeptidase 24.11 only partially contributes to this cleavage; (3) there exists in rat brain synaptic membranes a peptidase different from proline endopeptidase and endopeptidase 24.11 that is mainly responsible for inactivating neurotensin by cleaving at the Pro10-Tyr11 bond.     

Attenuation of pressor responses to intracerebroventricular angiotensin I by angiotensin converting enzyme inhibitors and their effects on systemic blood pressure in conscious rats

The components of the renin-angiotensin system exist in the brain but their physiological role is uncertain. The effects of two angiotensin converting enzyme (ACE) inhibitors, MK 421 (or its diacid) and captopril, on brain ACE activity, as measured by inhibition of the pressor response to intracerebroventricularly (i.c.v.) administered angiotensin I (AI), and the potential contribution of the central nervous system to their antihypertensive activity were evaluated in the present series of experiments. The diacid of MK 421 (1 and 10 ug) and captopril (3 and 10 ug) given i.c.v. to conscious normotensive rats reduced the pressor response to i.c.v. AI indicating that they can inhibit brain ACE. Responses to AII were unaffected. Oral administration of maximal antihypertensive doses of MK 421 (10 mg/kg) and of captopril (30 mg/kg) to normotensive rats did not attenuate pressor responses to i.c.v. AI indicating that brain ACE was not inhibited under these circumstances. Intracerebroventricular administration of MK 421 diacid, (10 and 30 ug) and captopril (30 and 100 ug) did not lower baseline blood pressure of spontaneously hypertensive rats. These experiments indicate that MK 421 and captopril can inhibit brain ACE but that the central renin-angiotensin system probably does not contribute to their antihypertensive activity.     

Hydrolysis of substance p and neurotensin by converting enzyme and neutral endopeptidase

Angiotensin I converting enzyme (ACE) and neutral endopeptidase ("enkephalinase"; NEP), were purified to homogeneity from human kidney. NEP cleaved substance P (SP) at Gln6-Phe7,-Phe8, and Gly9-Leu10 and neurotensin (NT) at Pro10-Tyr11 and Tyr11-Ile12. NEP hydrolyzed 0.1 mM SP, NT and their C-terminal fragments at the following rates (mumol/min/mg): SP1-11 = 7.8, SP4-11 = 11.7, SP5-11 = 15.4, SP6-11 = 15.6, SP8-11 = 6.7, NT1-13 = 2.9, and NT8-13 = 4.0. Purified ACE rapidly inactivated SP as measured in bioassay. HPLC analysis showed that ACE cleaved SP at Phe8-Gly9 and Gly9-Leu10 to release C-terminal tri- and dipeptide (ratio = 4:1). The hydrolysis was Cl- dependent and inhibited by captopril. ACE released mainly C-terminal tripeptide from SP methyl ester, but only dipeptide from SP free acid. Modification of arginine residues in ACE with cyclohexanedione or butanedione similarly inhibited hydrolysis of SP, bradykinin and Bz-Gly-Phe-Arg (80-93%) indicating an active site arginine is required for hydrolysis of SP. ACE hydrolyzed NT at Tyr11-Ile12 to release Ile12-Leu13. SP, NT and their derivatives (0.1 mM) were cleaved by ACE at the following rates (mumol/min/mg): SP1-11 = 1.2, SP methyl ester = 0.7, SP free acid = 8.5, SP4-11 = 2.4, SP5-11 = 0.9, SP6-11 = 1.4, SP8-11 = 0, NT1-13 = 0.2, and NT8-13 = 1.3. Peptide substrates were used as inhibitors of ACE (substrate = FA-Phe-Gly-Gly) and NEP (substrate = Leu5-enkephalin).(ABSTRACT TRUNCATED AT 250 WORDS)     

Suppression of plasma aldosterone by chronic ACTH administration is offset by converting enzyme inhibitor

In order to elucidate the mechanism of suppression of plasma aldosterone by chronic ACTH administration, especially the role of the renin-angiotensin system and dopamine, we administered ACTH with or without MK422, a converting enzyme inhibitor, to reduce the endogenous angiotensin II in rats, and measured the plasma renin activity, plasma corticoid concentrations and urinary dopamine excretion. The plasma aldosterone concentration (PAC) was decreased after chronic ACTH administration. However, in the ACTH + MK422 administered group, aldosterone suppression was not observed. It appeared therefore that the aldosterone suppressing mechanism was independent of the weakened renin-angiotensin system following chronic ACTH administration, since PAC was not decreased in the ACTH + MK422 administered group when angiotensin II might be completely eliminated. The urinary excretion of dopamine was significantly increased in the chronic ACTH + MK422 administered group as well as in the chronic ACTH administered group. This suggested that the inhibitory effect of dopamine on aldosterone did not contribute significantly to the suppression of plasma aldosterone. The present results suggest therefore that the mechanism of suppression of plasma aldosterone following chronic ACTH administration was not dependent on the renin-angiotensin system and dopamine.     

Determination of angiotensin-converting enzyme activity using dansyl-Phe-Ala-Arg as the substrate

Modificated method for the determination of the angiotensin converting enzyme (EC 3.4.15.1) activity in neural tissue is proposed. The methods is based on fluorimetrically determination of released dansyl-Phe from dansyl-Phe-Ala-Arg at pH 7.6. Km is 50 +/- 10 MM. The high specificity of the method is provide by using captopril, the high specific angiotensin converting enzyme inhibitor. The sensitivity of the method is 0.001 nmol/min per mg protein that it is lesser in 15-fold than the lowest enzyme activity in brain region. The distribution of the angiotensin converting enzyme activity that using the method in brain regions and peripherial tissues of rat is presented.     

Effect of captopril (SQ 14225) on blood pressure,plasma renin activity and angiotensin I converting enzyme activity.

Seven patients with essential hypertension and seven patients with hypertension associated with renal artery stenosis received captopril (SQ 14225), an inhibitor of angiotensin I converting enzyme. There was a significant reduction in mean blood pressure, from 176/113 +/- 4/3 mm Hg during the control period to 140/90 +/- 5/3 mm Hg during captopril administration. Five patients received captopril alone and nine patients needed hydrochlorothiazide in addition to control their blood pressure. Captopril produced a significant increase in peripheral plasma renin activity. When measured 12 hours after the administration of captopril the angiotensin I converting enzyme activity was found to be similar to that during the control period even though the blood pressure was at or near normal. These findings indicate that although captopril is an effective antihypertensive agent, its action does not depend only on inhibition of plasma angiotensin I converting enzyme activity.     

Angiotensin converting enzyme (ACE) and neprilysin hydrolyze neuropeptides: a brief history, the beginning and follow-ups to early studies

Our investigations started when synthetic bradykinin became available and we could characterize two enzymes that cleaved it: kininase I or plasma carboxypeptidase N and kininase II, a peptidyl dipeptide hydrolase that we later found to be identical with the angiotensin I converting enzyme (ACE). When we noticed that ACE can cleave peptides without a free C-terminal carboxyl group (e.g., with a C-terminal nitrobenzylamine), we investigated inactivation of substance P, which has a C-terminal Met(11)-NH(2). The studies were extended to the hydrolysis of the neuropeptide, neurotensin and to compare hydrolysis of the same peptides by neprilysin (neutral endopeptidase 24.11, CD10, NEP). Our publication in 1984 dealt with ACE and NEP purified to homogeneity from human kidney. NEP cleaved substance P (SP) at Gln(6)-Phe(7), Phe(7)[see text]-Phe(8), and Gly(9)-Leu(10) and neurotensin (NT) at Pro(10)-Tyr(11) and Tyr(11)-Ile(12). Purified ACE also rapidly inactivated SP as measured in bioassay. HPLC analysis showed that ACE cleaved SP at Phe(8)-Gly(9) and Gly(9)-Leu(10) to release C-terminal tri- and dipeptide (ratio = 4:1). The hydrolysis was Cl(-) dependent and inhibited by captopril. ACE released only dipeptide from SP free acid. ACE hydrolyzed NT at Tyr(11)-Ile(12) to release Ile(12)-Leu(13). Then peptide substrates were used to inhibit ACE hydrolyzing Fa-Phe-Gly-Gly and NEP cleaving Leu(5)-enkephalin. The K(i) values in microM were as follows: for ACE, bradykinin = 0.4, angiotensin I = 4, SP = 25, SP free acid = 2, NT = 14, and Met(5)-enkephalin = 450, and for NEP, bradykinin = 162, angiotensin I = 36, SP = 190, NT = 39, Met(5)-enkephalin = 22. These studies showed that ACE and NEP, two enzymes widely distributed in the body, are involved in the metabolism of SP and NT. Below we briefly survey how NEP and ACE in two decades have gained the reputation as very important factors in health and disease. This is due to the discovery of more endogenous substrates of the enzymes and to the very broad and beneficial therapeutic applications of ACE inhibitors.     

Tubuloglomerular feedback-dependent influence of angiotensin II on the kidney in rats

To examine the modulatory role of angiotensin II on the tubuloglomerular feedback (TGF) mechanism, TGF responses were assessed during control conditions, converting enzyme inhibition (CEI; MK 422, 0.6 mg/kg.hr) and during continued CEI with the replacement of angiotensin II. TGF responses were assessed from stop flow pressure (SFP) feedback responses obtained during step increases in the late proximal perfusion rate from 0-40 nl/min. SFP values in the absence of perfusion were used to estimate glomerular pressure (GP) under conditions where the influence of the TGF mechanism should be at a minimum. During CEI, the arterial pressure decreased from 124 +/- 3 to 106 +/- 3 mmHg and the estimated GP decreased from 53 +/- 1.4 to 49 +/- 0.8 mmHg. There was a marked attenuation in the magnitude of SFP feedback responses from 11.0 +/- 1.3 to 2.7 +/- 0.6 mmHg. TGF feedback responses, however, were restored towards normal during superimposed angiotensin II infusion (7.7 +/- 0.9 mmHg). These results indicate that converting enzyme inhibition decreases the effects of angiotensin II on the kidney through TGF dependent mechanism.     

Novel substrates for angiotensin I converting enzyme

Homogenous human angiotensin converting enzyme (EC 3.4.15.1) cleaves dipeptides from the C-terminus of substrates containing a free carboxyl group. In this study we demonstrate that peptides containing a C-terminal nitrobenzylamine are also cleaved by the enzyme. The hydrolysis of these substrates is inhibited by the specific converting enzyme inhibitors captopril and MK421 as well as by anti-converting enzyme antibody. Sodium chloride accelerates the rate of hydrolysis forty-fold. The product of the reaction, an amino acid nitrobenzylamide, was identified by thin layer chromatography and high performance liquid chromatography. These results suggest that the carboxyl group is not an absolute requirement for substrate hydrolysis.     

Preventive effect of angiotensin I on weight reduction in the adrenal glands of DOCA/salt hypertensive rats

We examined the effect of angiotensin I (AI), without the effect of angiotensin II (AII) converted from AI, on the weight of the adrenal glands, adrenal corticosterone (B) and adrenal aldosterone under conditions where the renin-angiotensin system was suppressed, since a reduction in the size of the adrenal glands is often observed in DOCA/salt hypertensive rats. Sixty male Wistar rats fed on a 1% NaCl solution were divided into 6 groups as follows: a) Salt group: received sesame oil and vehicle, b) Salt + C group: received sesame oil and MK422 (0.14 mg/day), an angiotensin converting enzyme inhibitor (CEI), c) DOCA group: received DOCA (30 mg/week) and vehicle, d) DOCA + A group: received DOCA and AI (0.5 mg/kg/day), e) DOCA + A + C group: received DOCA and AI with MK422, and f) DOCA + C group: received DOCA and MK422. After 4 weeks, the rats were sacrificed to sample their blood and remove their adrenal glands. There was no significant difference in adrenal B among the groups apart from the DOCA + C group. Adrenal aldosterone was lower in the groups of DOCA/salt hypertensive rats than in the Salt group and Salt + C group. Furthermore, the DOCA + A + C group and DOCA + C group had lower adrenal aldosterone levels than the DOCA group and DOCA + A group.(ABSTRACT TRUNCATED AT 250 WORDS)     

Regional Metabolism of Met-Enkephalin and Cholecystokinin on Intact Rat Brain Slices: Characterization of Specific Peptidases

Abstract: The metabolism of Met-enkephalin and cholecystokinin (CCK) 8-(sulfated) by intact microslices was studied in rat brain regions. Incubation of brain slices with Met-enkephalin (400 µ M ) resulted in a linear rate of disappearance of parent peptide and appearance of metabolic fragments whose rate of accumulation was specific to brain region. The degradative rate (pmol/min/mg of protein) of Met-enkephalin was high in caudate-putamen (5,160 ± 120) and lower in nucleus accumbens (3,630 ± 110) and frontal cortex (3,180 ± 120). Inhibition of aminopeptidases decreased Met-enkephalin degradation (50–97% vs. control) in frontal cortex but was less effective in caudate-putamen (20–34%). Tyr-Gly-Gly and Phe-Met were recovered in caudate-putamen and nucleus accumbens, whereas negligible quantities of these fragments were recovered from frontal cortex. Phosphoramidon, an inhibitor of neutral endopeptidase 24.11, decreased Met-enkephalin degradation in caudate-putamen (14%) but had no effect on that in frontal cortex. A cocktail of bestatin or leuhistin (inhibitors of aminopeptidases), phosphoramidon, and captopril (an inhibitor of angiotensin converting enzyme) protected Met-enkephalin from degradation (recovery >95%) in caudate-putamen. CCK 8-(sulfated) degradation on slices from caudate-putamen, nucleus accumbens, and frontal cortex was not altered by inhibitors of neutral endopeptidase 24.11, metalloendopeptidase 24.15, angiotensin converting enzyme, or thiol proteases. Inhibitors of either aminopeptidases or serine proteases produced small reductions (13–30%) in CCK degradation in each region. These data provide evidence for regional and structural specificity in terminating the actions of neuropeptides.     

Low molecular weight angiotensin I converting enzyme from rat lung.

A low molecular weight angiotensin I converting enzyme (light angiotensin enzyme) was isolated from a homogenate of rat lung subjected to dialysis against sodium acetate at pH 4.8. This enzyme has a molecular weight of 84 000 on Sephadex G-200 and a molecular weight of 91 000 on SDS-poly-acrylamide gel as compared with a molecular weight of 139 000 for angiotensin I converting enzyme on SDS-polyacrylamide. Light angiotensin enzyme was activated by NaCl and inhibited by EDTA, angiotensin II, and bradykinin potentiating factor nonapeptide. Light angiotensin enzyme cross-reacted with antibody prepared against angiotensin I converting enzyme and stained with periodic acid-Schiff reagent as a glycoprotein. The evidence suggests that light angiotensin enzyme is a fragment of the higher molecular weight enzyme.