Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: CAS:1160-54-9 (Z-Gly-Pro)
 78 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The endopeptidase, post-proline cleaving enzyme, has been purified 10,500-fold in an overall yield of 18% from lamb kidney. The enzyme possesses a specific activity of 45 mumol/mg/min as tested with the substrate Z-Gly-Pro-Leu-Gly (Km = 6.0 X 10(-5)), has a molecular weight of 115,000, is comprised of two subunits with a molecular weight of 57,000, and exhibits maximal activity at pH 7.5 to 8.0. With the exception of the -Pro-Pro linkage, the -Pro-X-peptide bond (X equals L- and D-amino acid residues) located internally in the peptide sequence can be hydrolyzed (cleavage occurs faster when X = lipophilic side chain as compared to X = acidic side chain). The appropriate -Pro-X- bonds in zinc-free porcine insulin, oxytocin, arginine vasopressin, angiotensin II, bradykinin-potentiating factor were cleaved. Human gastrin, adrenocorticotropic hormone, denatured guinea pig skin collagen, and ascaris cuticle collagen were not degraded. Dipeptides with the structure Z-Pro-LD-X competitively inhibit post-proline cleaving enzyme.
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PMID:Post-proline cleaving enzyme. Purification of this endopeptidase by affinity chromatography. 1 73

Synthesis and application of the first fluorogenic substrate, N-carbobenzoxyglycylprolyl-4-methylcoumarinyl amide (Z-Gly-Pro-MeCouNH) for the determination of the post-proline cleaving enzyme (EC 3.4.21.-) were reported. Maximal activity of the enzyme purified from lamb kidney for the new substrate was observed at pH 7.0. This substrate showed a higher affinity (Km = 0.02 mM) for the enzyme than the proline containing substrates studied previously and allowed the detection of 10-50 ng post-proline cleaving enzyme activity per ml sample after a 1 min incubation period. Distribution of post-proline cleaving enzyme and other proline specific peptidases in rat tissues was studied using Z-Gly-Pro-MeCouNH and other proline-containing substrates. High post-proline cleaving enzyme activity was observed in testis, liver and skeletal muscle. Inhibition experiments indicated that post-proline cleaving enzyme activity was completely inactivated by 0.1 mM diisopropylphosphofluoridate and Z-Gly-Pro-chloromethylketone, as had been found in the case of the enzyme isolated from lamb kidney. Activity in human body fluids was also tested for levels of post-proline cleaving enzyme activity using Z-Gly-Pro-MeCouNH and semen was found to show the highest cleaving activity.
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PMID:Post-proline cleaving enzyme. Synthesis of a new fluorogenic substrate and distribution of the endopeptidase in rat tissues and body fluids of man. 47 24

High prolyl endopeptidase (post-proline cleaving enzyme) [EC 3.4.21.26] activity was detected in fruit bodies of shakashimeji (Lyophyllum cinerascens), tsukuritake (mushroom: Agaricus bisporus), hirohachichitake (Lactarius hygrophoroides), and yaburebenitake (Russula lepida) which belong to the genus Basidiomycetes. Cell-free extract of shakashimeji showed high activities of proline iminopeptidase and arylamidase as well as prolyl endopeptidase. The prolyl endopeptidase was purified from the extract of shakashimeji by sequential chromatographies on DEAE-Toyopearl, DEAE-Sephadex and hydroxyapatite, and high-performance liquid chromatography with a DEAE-5PW column. The purified enzyme was homogeneous as judged by disc gel electrophoresis. The enzyme was most active at pH 6.8 as checked with Z-Gly-Pro-beta-naphthylamide as a substrate and was stable in the range of pH 5.8-7.4. The isoelectric point of the enzyme was 5.2 and the molecular weight was estimated to be 76,000 by gel filtration on Sephadex G-150 and by sodium dodecyl sulfate (SDS) gel electrophoresis, suggesting that the enzyme was a monomer. The enzyme was completely inhibited by diisopropyl fluorophosphate (DFP), Z-Gly-Pro-CH2Cl, and Z-Pro-prolinal, while it was not inhibited by p-chloromercuribenzoate (PCMB), phenylmethylsulfonyl fluoride (PMSF), or metal chelators. It was estimated that at least five subsites were concerned with the enzyme-substrate binding. Among them, the S1, S2, and S1' sites showed high stereospecificity, as in mammalian, microbial, and plant enzymes. The enzyme hydrolyzed TRH at the carboxyl side of the proline residue. The mushroom enzyme, that was sensitive to DFP, Z-Pro-prolinal, and Z-Gly-Pro-CH2Cl, but not to PCMB, were quite similar in characteristics to the Flavobacterium enzyme.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Studies on prolyl endopeptidase from shakashimeji (Lyophyllum cinerascens): purification and enzymatic properties. 307 34

Post-proline cleaving enzyme (PPCE; EC 3.4.21.26) is a proline specific endopeptidase capable of hydrolyzing biologically active peptides. The present studies examined the hydrolysis of kinin- and angiotensin-related peptides by cytosolic PPCE purified from porcine kidney. PPCE hydrolysis of the synthetic substrate Z-Gly-Pro-MCA (30.7 +/- 0.3 mumol . min-1 . mg-1) was competitively inhibited by saralasin, bradykinin, des(Arg9)bradykinin, [Leu8], des(Arg9)bradykinin and angiotensin II (IC50 = 0.5 to 7.0 microM). Qualitative TLC studies demonstrated that each peptide was degraded by hydrolysis on the carboxyl side of proline residues (positions 7 or 8). Quantitative HPLC studies established that peptide degradation was optimal at pH 8.2 to 8.7 and was inhibited by the specific PPCE inhibitor Z-Pro-prolinal (IC50 = 0.8 +/- 0.1 nM). Conversely, degradation was unaffected by inhibitors of aminopeptidases (amastatin), neutral endopeptidase (phosphoramidon), carboxypeptidase N (MERGETPA) or angiotensin I converting enzyme (captopril). Apparent Km values, obtained from Lineweaver-Burk analysis, were comparable for all kinin and angiotensin peptides (Km = 5.5 to 12.8 microM), whereas Vmax values ranged from 1.7 mumol . min-1 . mg-1 for angiotensin II to 0.44 mumol . min-1 . mg-1 for saralasin. These data are consistent with a role for PPCE in the degradation of kinins and angiotensin in vivo.
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PMID:Kinin and angiotensin metabolism by purified renal post-proline cleaving enzyme. 347 49

Post-proline cleaving enzyme (PPCE) was purified from porcine kidney cytosol. The purified enzyme bound [125I-Tyr5]-bradykinin but neither [125I-Tyr1]-kallidin nor [125I-Tyr8]-bradykinin. Scatchard analysis of the data was consistent with a single class of binding sites with a Kassoc = 1.3 +/- 0.1 X 10(8) M-1. The optimal pH for [125I-Tyr5]-bradykinin binding was 6.8. The specificity of binding was evaluated with sixty-seven bradykinin analogs. The catalytic activity of the enzyme was measured with N-benzyloxycarbonyl-Gly-Pro-methylcoumarinyl-7-amide (Z-Gly-Pro-MCA). The optimal pH for hydrolysis of this substrate was broad and centered at 8.3. The apparent Km and Vmax were obtained from Lineweaver and Burk plots and were 4.8 +/- 0.4 X 10(-5) M and 42 +/- 5 mumoles X mg-1 X min-1 respectively. The IC50 values for bradykinin, diisopropylfluorophosphate (DFP), and N-benzyloxycarbonyl-Pro-Prolinal (Z-Pro-Prolinal) to inhibit Z-Gly-Pro-MCA hydrolysis by PPCE were 5.9 +/- 1.4 X 10(-7) M, 8.8 +/- 3.1 X 10(-7) and 7.9 +/- 0.3 X 10(-9) M respectively. Corresponding values for inhibition of [125I-Tyr5]-bradykinin binding by PPCE were 5.1 +/- 2.3 X 10(-9) M, 1.2 +/- 0.3 X 10(-6) M and 1.4 +/- 0.6 X 10(-8) M.
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PMID:Specific, high-affinity bradykinin binding by purified porcine kidney post-proline cleaving enzyme. 354 40

Post-proline endopeptidase (PPE, EC 3.4.21.26) was purified 3,450 times from human lung. PPE was routinely assayed with the artificial substrate, carbobenzoxy-glycyl-L-prolyl-p-nitroanilide (Z-Gly-Pro-pNA). The pH optimum was 7.4, and the Mr was 77,000. Thiol blocking agents were strongly inhibitory but serine blocking agents were not inhibitory. No metal ions were required for activity, but heavy metal ions such as Hg2+, Cu2+, Cd2+, and Zn2+ completely inactivated the enzyme. Both dithiothreitol (DTT) and ethylenediaminetetraacetic acid (EDTA) were required to stabilize PPE activity. Michaelis constant values for Z-Gly-Pro-pNA and carbobenzoxy-glycyl-L-prolyl-2-naphthylamide were 0.36 and 0.10 mmol/l, respectively. PPE cleaved vasoactive peptides including bradykinin (BK) and des-(Arg9)-BK (Pro3-Gly4 and Pro7-Phe8 bonds), angiotensins I and II (Pro7-Phe8 bond), substance P (Pro4-Gln5 bond), and oxytocin (Pro7-Leu8 bond). Each of these peptides inhibited PPE-catalyzed hydrolysis of Z-Gly-Pro-pNA competitively. BK had the lowest Ki value (2.35 mumol/l) and oxytocin had the highest Ki value (84.0 mumol/l). PPE was not inhibited by captopril, a potent inhibitor of angiotensin converting enzyme, which also cleaves the Pro7-Phe8 bond of BK.
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PMID:Human lung post-proline endopeptidase: purification and action on vasoactive peptides. 354 26

An inhibitor(s) for post-proline cleaving enzyme was checked by using a fluorogenic substrate, Z-Gly-Pro-4-methyl coumarinamide, and was found to be distributed widely in rat and porcine organs. The highest inhibitory activity on the enzyme was observed in pancreas and the inhibitor was partially purified from porcine pancreas extract by heat treatment, chromatographies on DEAE-Sephadex and Sephadex G-50 and affinity chromatography on trypsin-Sepharose. This inhibitor was very stable against temperature, pH and trichloroacetic acid treatment. The molecular weight was estimated to be 6500 by gel filtration. This inhibitor was highly specific for prolyl endopeptidases from mammals and Flavobacterium and inhibited the enzyme competitively. It acted neither on proline specific exopeptidases such as dipeptidyl aminopeptidase IV, proline aminopeptidase, prolidase, nor usual endopeptidases such as trypsin and alpha-chymotrypsin.
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PMID:An inhibitor for post-proline cleaving enzyme; distribution and partial purification from porcine pancreas. 618 66

A post-proline cleaving enzyme and its endogenous inhibitor have been demonstrated to be present in sperm of the ascidian, Halocynthia roretzi. The enzyme was extracted with artificial sea water from frozen and thawed sperm and isolated from accompanying acrosin-like and chymotrypsin-like enzymes by DEAE-cellulose chromatography. It was then separated from the endogenous inhibitor by ammonium sulfate fractionation and DEAE-Sephacel chromatography. Three subsequent chromatographic operations using hydroxylapatite, Sephadex G-150 and Z-Gly-Pro-Leu-Gly-aminohexyl-Sepharose yielded the highly purified enzyme. The molecular weight and isoelectric point of the enzyme were estimated to be 66,000 and 5.5, respectively. The pH optimum of the activity was 7.0. The enzyme was inactivated with diisopropylphosphorofluoridate, phenylmethylsulfonyl fluoride, Z-Gly-Pro-chloromethyl ketone and sulfhydryl-directed reagents; these inhibitor susceptibilities were similar to those reported for the enzymes of mammalian origins. The ascidian enzyme hydrolyzed oxytocin, angiotensin II, luteinizing hormone releasing hormone and neurotensin at the carboxyl side of proline residues. The endogenous inhibitor was heat stable. The molecular weight of its main component was estimated to be about 8,000. The presence of salt at high concentrations weakened the enzyme-inhibitor interaction. Z-Gly-Pro-chloromethyl ketone inhibited fertilization of the ascidian, suggesting possible involvement of the post-proline cleaving enzyme in fertilization.
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PMID:Isolation and characterization of a post-proline cleaving enzyme and its inhibitor from sperm of the ascidian, Halocynthia roretzi. 636 Oct 7

A post-proline cleaving enzyme [prolyl endopeptidase, EC 3.4.21.26] was purified about 3,700-fold from an extract of bovine brain by a series of column chromatographies on DEAE-Sephadex, hydroxyapatite and PCMB-T-Sepharose, and gel filtration on Sephadex G-200 using N-carbobenzoxy-Gly-Pro-beta-naphthylamide (Z-Gly-Pro-2-NNap), thyrotropin releasing hormone (TRH) and oxytocin as substrates. The purified enzyme appeared homogeneous as judged by disc gel and SDS gel electrophoreses. The enzyme was most active at pH 7.5 and 7.2 with Z-Gly-Pro-2-NNap and TRH, respectively, and hydrolyzed peptide bonds involving Pro-X (X=amino acid, peptide, ester and amide) bonds of synthetic substrates, oxytocin, vasopressin, neurotensin, substance P, tuftsin, bradykinin, and insulin B chain. However, the enzyme was inert toward collagen, gelatin, and casein. The enzyme was completely inactivated by diisopropylphosphorofluoridate (DFP), Z-Gly-Pro-chloromethyl ketone and p-chloromercuribenzoate (PCMB), while it was not inhibited by phenylmethane sulfonylfluoride (PMSF) or metal chelators. Determination of the amino acid composition revealed that the enzyme contained 25 half-cystines. Modification of three cysteine residues of the enzyme by PCMB led to complete inactivation. The isoelectric point of the enzyme was 4.8, and the molecular weight was estimated to be 76,000 by ultracentrifugal analysis and 75,000-74,000 by both gel filtration and sodium dodecyl sulfate (SDS) gel electrophoresis, suggesting that the enzyme is present as a monomer. These results indicate that the post-proline cleaving enzyme from bovine brain is very similar to those previously purified from lamb brain and kidney in its enzymatic properties, substrate specificity and physicochemical properties, in sharp contrast with the results obtained by Tate, who reported that the bovine brain prolyl endopeptidase was inert toward oxytocin, vasopressin and bradykinin.
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PMID:Post-proline cleaving enzyme (prolyl endopeptidase) from bovine brain. 636 Oct 10

A post-proline cleaving enzyme [post-proline endopeptidase: EC 3.4.21.26] was purified from lamb brain by a series of column chromatographies on DEAE-Sephadex, hydroxyapatite and Sephadex G-150. The purified enzyme appeared homogeneous on disc gel and sodium dodecyl sulfate (SDS) gel electrophoreses. The enzyme was most active at pH 7.0 with carbobenzoxy-Gly-Pro-beta-naphthylamide (Z-Gly-Pro-2-NNap) as a substrate and catalyzed the hydrolysis of oxytocin, vasopressin, thyrotropin releasing hormone (TRH), substance P, luteinizing hormone releasing hormone (LH-RH), and angiotensin at the carboxyl side of their proline residues, except for the Pro2-Lys3 bond in substance P. From the results of subsite mapping using synthetic peptides, five subsites, S3 to S2', for substrate interaction with the enzyme were deduced to be present, and high stereospecificity was observed at S2, S1, and S1'. The isoelectric point of the enzyme was at pH 4.9, and the molecular weights estimated by gel filtration and SDS gel electrophoresis were 74,000 and 77,000, respectively. The enzyme was markedly inhibited by diisopropylphosphoro fluoridate (DFP), carbobenzoxy-Gly-Pro-chloromethyl ketone (Z-Gly-Pro-CH2Cl), p-chloromercuribenzoate (PCMB), Hg2+, and Cu2+ ions. These enzymatic and protein chemical properties of post-proline cleaving enzyme from lamb brain closely resemble those of the lamb kidney enzyme, except for the molecular weight. In the present work, however, we decided that the molecular weight of the enzyme from lamb kidney was also 74,000, which is different from that reported previously (J. Biol. Chem. 251, 7593 (1976) but is in accord with the value of post-proline cleaving enzyme from lamb brain.
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PMID:Post-proline cleaving enzyme from lamb brain. 702 30


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