Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.15.1 (ACE)
18,300 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Stopped-flow radiationless energy-transfer kinetics have been used to examine the effects of chloride on the hydrolysis of Dns-Lys-Phe-Ala-Arg by angiotensin converting enzyme. The kinetic constants for hydrolysis at pH 7.5 and 22 degrees C in the presence of 300 mM sodium chloride were KM = 28 microM and kcat = 110 s-1, and in its absence, KM = 240 microM and kcat = 68 s-1. The apparent binding constant for chloride was 4 mM, and the extent of chloride activation in terms of kcat/KM was 14-fold. The effects of chloride on the pre-steady-state were examined at 2 degrees C. In the presence of chloride, two distinct enzyme-substrate complexes were observed, suggesting multiple steps in substrate binding. The initial complex was formed during the mixing period (kobsd greater than 200 s-1) while the second complex was formed much more slowly (kobsd = 40 s-1 when [S] = 5 microM and [NaCl] = 150 mM). Strikingly, in the absence of chloride, only a single, rapidly formed enzyme-substrate complex was observed. These results are consistent with a nonessential activator kinetic mechanism in which the slow step reflects conversion of an initially formed complex, (E X Cl- X S)1, to a more tightly bound complex, (E X Cl- X S)2.
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PMID:Observation of a chloride-dependent intermediate during catalysis by angiotensin converting enzyme using radiationless energy transfer. 303 49

Carboxypeptidase N (CPN, kininase I) and kininase II (angiotensin converting enzyme) activities were measured simultaneously in blood plasma and synovial fluid in patients suffering from rheumatoid arthritis (RA), psoriatic arthritis (PA) and osteoarthritis (OA) and in the plasma of normal volunteers. CPN levels (defined as the rate of hydrolysis of furylacryloyl-Ala-Lys) in blood were modestly increased and correlated with erythrocyte sedimentation rate in RA and PA. Based on the hydrolysis of synthetic substrates, CPN activity was much higher than kininase II activity in synovial fluid (SF). SF kininase activities were always inferior to the blood levels in all patients and were correlated with the logarithm of SF leukocyte counts, an indicator of the intensity of inflammation. In addition, CPN and albumin levels in SF were highly correlated when expressed as a percent of the plasma concentrations. Biochemical properties of CPN in crude SF confirmed its similarity to blood CPN. Polymorphonuclear leukocytes derived from inflammatory SF did not release CPN. It is concluded that kininases diffuse from the blood into SF through increased vascular permeability and that CPN could be a major metabolic pathway for kinins in this form of exudate. CPN leads to the formation of des-Arg kinins, selective agonists of the B1 receptors for kinins.
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PMID:Carboxypeptidase N (kininase I) activity in blood and synovial fluid from patients with arthritis. 304 Nov 37

Dipeptidase (dipeptide hydrolase [EC 3.4.13.11]) has been purified to homogeneity and crystallized from the cell extract of Bacillus stearothermophilus IFO 12983. The enzyme has a molecular weight of about 86,000, and is composed of two subunits identical in molecular weight (43,000). The enzyme contains 2 g atoms of zinc per mol of protein. A variety of dipeptides consisting of glycine or only L-amino acids serve as substrates of the enzyme; Km and Vmax values for L-valyl-L-alanine are 0.5 mM and 68.0 units/mg protein, respectively. The enzyme is significantly stable not only at high temperatures but also on treatment with protein denaturants such as urea and guanidine hydrochloride. The enzyme also catalyzes hydrolysis of several N-acylamino acids with Vmax values 3-30% of those for the hydrolysis of dipeptides. The thermostable dipeptidase shares various properties with bacterial aminoacylase [EC 3.5.1.14]: their subunit molecular weight, metal content and requirement, amino acid composition, and amino acid sequence in the N-terminal region are very similar.
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PMID:Thermostable dipeptidase from Bacillus stearothermophilus: its purification, characterization, and comparison with aminoacylase. 313 45

The design rationale for a new series of tripeptide derived angiotensin converting enzyme (ACE) inhibitors, which we term "ketomethylureas", is described. Analogs of tripeptide substrates (i.e. N-benzoyl-Phe-Ala-Pro) in which the nitrogen atom of the scissile amide bond and the adjacent asymmetric carbon atom of the penultimate amino acid residue are formally transposed give rise to this novel class of inhibitors. The most potent ketomethylureas inhibit ACE with I50 values in the nM range.
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PMID:Ketomethylureas. A new class of angiotensin converting enzyme inhibitors. 323 70

The concentration of luteinizing hormone releasing hormone (LHRH) (pGlu-His-Trp-Ser-Tyr-Gly-Leu-Arg-Pro-Gly-NH2), which reaches the anterior pituitary via the hypothalamo-hypophyseal portal system, appears to be controlled in part by the rate of LHRH degradation within the hypothalamus and/or pituitary. Specific, active site-directed endopeptidase inhibitors synthesized in our laboratory were used to identify the enzyme(s) involved in LHRH degradation by hypothalamic and pituitary membrane preparations, and by an intact anterior pituitary tumor cell line (AtT20). Incubation of LHRH with pituitary and hypothalamic membrane preparations led to the formation of pGlu-His-Trp (LHRH1-3) as the main reaction product. Under the same conditions, addition to the incubation mixtures of captopril, an inhibitor of the angiotensin converting enzyme, led to accumulation of pGlu-His-Trp-Ser-Tyr (LHRH1-5) and, to a lesser extent, pGlu-His-Trp-Ser-Tyr (LHRH1-6). The degradation of LHRH and the formation of the N-terminal tri- and pentapeptides was blocked by N-[1-(R,S)-carboxy-3-phenylpropyl]-Ala-Ala-Phe-p-aminobenzoate (cFP-AAF-pAB), a specific, active site directed inhibitor of endopeptidase-24.15. Some inhibition of LHRH degradation and formation of the N-terminal hexapeptide was also obtained in the presence of N-[1-carboxy-2-phenylethyl]-Phe-p-aminobenzoate (cFE-F-pAB), an inhibitor of endopeptidase-24.11. Similar results were obtained with AtT20 cell membranes and with intact AtT20 cells in monolayer culture. Following cleavage by endopeptidases the C-terminal part of LHRH was rapidly degraded by aminopeptidases. Superactive analogs of LHRH in which Gly6 was replaced by a D-amino acid are resistant to degradation by both endopeptidase-24.11 and -24.15. In vivo, when LHRH was injected directly into the third ventricle of rats, the presence of cFP-AAF-pAB inhibited LHRH degradation. It is concluded that LHRH degradation is primarily initiated by the membrane-bound form of endopeptidase-24.15 to yield pGlu-His-Trp-Ser-Tyr and to a lesser extent by endopeptidase-24.11 to yield pGlu-His-Trp-Ser-Tyr-Gly.
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PMID:Endopeptidase-24.15 is the primary enzyme that degrades luteinizing hormone releasing hormone both in vitro and in vivo. 329 5

Cathepsin B from brain exhibited both endopeptidase and dipeptidyl carboxypeptidase activity. Recently the factors, contributing to dipeptidyl carboxypeptidase properties of brain cathepsin B, were identified: I. occupation of the enzyme S3 subsite, 2. free C-terminal group of the substrate, 3. specific interaction between the split off dipeptide and the enzyme active site. The identification was carried out using angiotensin I, its C-end tripeptide and chromophore oligopeptides containing p-nitrophenylalanine residue. C-terminal dipeptide was split off in the proopioid peptides dynorphins 1-7 and 1-8, Met-enkephalin-Arg6-Phe7, Met-enkephalin-Arg6-Gly7-Leu8; the enzyme hydrolyzed also the C-terminal dipeptide bond in Leu- and Met-enkephalins without the subsequent hydrolysis of the remaining tripeptide. D-Ala2, D-Leu5-enkephalin were not hydrolyzed; the bond Arg9-Pro10 was resistant to proteolysis in dynorphin 1-11. Cathepsin B split off the C-terminal dipeptide in synthetic substrates Leu-Trp-Met-Arg-Phe-Ala and Trp-Met-Arg-Phe-Ala but not in Met-Arg-Phe-Ala. These results 06.08 M-15 demonstrated the essential role of branched-chain amino acid residue at the position of P2 and/or P3 of substrates for the enzyme dipeptidyl carboxypeptidase activity. The data obtained suggest that Arg residue at the position P2 (dynorphin 1-7) slowed down, D-amino acid at the position P2 (D-Ala2, D-Leu5-enkephalin) and Pro-Lys bond at the position P1-P2 (dynorphin 1-11) inhibited the cathepsin B dipeptidyl carboxypeptidase activity.
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PMID:[Brain cathepsin as dipeptidylcarboxypeptidase transforming provasopressor, pro-opioid and model peptides]. 331 15

The action of bovine spleen cathepsin B as a dipeptidyl carboxypeptidase on newly synthesized substrates of the type peptidyl-X-p-nitrophenylalanyl (Phe(NO2))-Y (X,Y = amino acid residue) or 5-dimethylaminonaphthalene-1-sulfonyl (Dns)-peptidyl-X-Phe(NO2)-Y was investigated. The kinetic parameters of hydrolysis of the X-Phe(NO2) bond were determined by difference spectrophotometry (delta epsilon 310 = 1600 M-1 cm-1) or by spectrofluorometry by following the five- to eightfold increase of Dns-group fluorescence with excitation at 350 nm and emission at 535 nm. The substrates were moderately sensitive to cathepsin B; kcat varied from 0.7 to 4 s-1 at pH 5 and 25 degrees C; Km varied from 6 to 240 microM. The very acidic optima of pH 4-5 are characteristic for dipeptidyl carboxypeptidase activity of cathepsin B. Bovine spleen cathepsins S and H had little and no activity, respectively, when assayed with Pro-Glu-Ala-Phe(NO2)-Gly. These peptides should be a valuable tool for routine assays and for mechanistic studies on cathepsin B.
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PMID:Chromophoric and fluorophoric peptide substrates cleaved through the dipeptidyl carboxypeptidase activity of cathepsin B. 331 52

In order to evaluate the structural/functional roles of Met residues in an octadecapeptide pigment-dispersing hormone (PDH: Asn-Ser-Gly-Met-Ile-Asn-Ser-Ile-Leu-Gly-Ile-Pro-Arg-Val-Met-Thr-Glu-Ala- NH2), first described as light-adapting distal retinal pigment hormone (DRPH) from Pandalus, three analogs were synthesized: Nle4-PDH, Nle15-PDH, and Nle4,15-PDH. When tested for melanophore pigment-dispersing activity in destalked Uca, all three Nle-analogs were more potent than unsubstituted PDH. Performic acid oxidation caused a marked loss of potency of PDH, Nle4-PDH, and Nle15-PDH. The analog Nle4,15-PDH was resistant to oxidation and displayed 6-fold higher potency than PDH. Thus Met4 and Met15 are not essential for the PDH activity. The oxidation-induced loss of activity of unsubstituted PDH may result from introduction of oxygen (in methionine sulfone) and a consequent conformational change in the octadecapeptide.
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PMID:Substitution of norleucine for methionine residues in a crustacean pigment-dispersing hormone. 384 Aug 88

This study deals with the effect of deamidation and C-terminal truncation on the potency of an octadecapeptide pigment-dispersing hormone (PDH: Asn-Ser-Gly-Met-Ile-Asn-Ser-Ile-Leu-Gly-Ile-Pro-Arg-Val-Met-Thr-Glu-Ala- NH2), first described as light-adapting distal retinal pigment hormone (DRPH) from Pandalus borealis. Bioassay of synthetic analogs for melanophore pigment dispersion in destalked fiddler crabs (Uca pugilator) showed that deamidation causes a 300-fold decrease in potency. The analogs 1-17 NH2 and 1-16 NH2 were about 3 times more potent than 1-18-OH. Further truncation led to decreases in potency, with the peptide 1-9-NH2 being the smallest C-terminal deletion analog to display activity (0.001% potency). Smaller analogs (1-8-NH2, 1-6-NH2 and 1-4-NH2) were inactive when tested in doses as high as 500 nmoles/crab. On the basis of our earlier work on N-terminal deletion analogs and the present findings the residues 6 to 9 seem to be important for PDH action.
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PMID:C-terminal deletion analogs of a crustacean pigment-dispersing hormone. 384 33

We have utilized a highly sensitive radiationless energy transfer (RET) assay to investigate the effect of anions on the activity of carboxypeptidase A (CPD-A). The RET kinetic method visualizes the ES complex directly and thus enables both the mode of action of anions and the quantitation of their effect to be determined at a single substrate concentration. In marked contrast to the activating effect of anions on the closely related metalloprotease, angiotensin converting enzyme, Cl-, and other anions inhibit CPD-A catalysis. NaCl inhibits the hydrolysis of Dns-Ala-Ala-Phe throughout the pH range 6-10. Other di- and tripeptides are similarly inhibited while their ester analogues are affected only slightly. Changes in the type of cation [e.g., Na+, Li+, K+, Ca2+, and (CH3)4N+] at a constant [Cl-1] of 0.1 M showed no difference in the extent of inhibition, whereas with anion substitution the differences were marked. In all cases, the inhibition was partially competitive. At pH 5.9, the Ki values for the free enzyme are 51 (Cl-), 17 (N3-), 2.1 (SO4(2-)), and 0.21 mM (H2PO4-), and for the ES complex, the KI' values are 1000, 720, 42, and 13 mM, respectively. The other anions were shown to act at the chloride site. The results indicate that investigations of anion inhibition in 1 M NaCl, a typical assay condition, may be greatly hindered by the presence of Cl-. Thus, the competitive binding mode of phenylacetate toward peptide hydrolysis is greatly decreased by the presence of 1 M Cl- ion while its noncompetitive component is unaffected.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Kinetic analysis by stopped-flow radiationless energy transfer studies: effect of anions on the activity of carboxypeptidase A. 395 97


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