Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.21.37 (neutrophil elastase)
 4,078 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Trifluoroacetylpeptide anilides are powerful reversible inhibitors of human neutrophil elastase (HNE), a serine protease implicated in the pathogenesis of pulmonary emphysema. The in vitro effectiveness of three inhibitors, CF3CO-Phe-Ala-NH-p-C6H4-CF3 (1), CF3CO-Val-Ala-NH-p-C6H4-CF3 (2) and CF3CO-Lys-Ala-NH-p-C6H4-CH(CH3)2 (3) was analyzed. The protection of lung tissue sections of rats from the degradation induced by HNE has been evaluated quantitatively by automated image analysis. Inhibitor 1 (22 microM), 2 (50 microM) or 3 (35 and 70 microM) significantly reduced the HNE-induced degradation of the elastin network by 75, 42, 54 and 44%, respectively. Inhibitor 3 was tested intratracheally on an experimental model of pulmonary emphysema. Rats that received the elastase inhibitor 1 h before instillation of HNE were significantly protected by 40% from experimental emphysema. Reduced protections were observed with the treatment by the inhibitor 1 or 4 h after challenge with the enzyme.
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PMID:Protection of rat lung from elastase-induced elastic fiber degradation in vitro and from emphysema in vivo by a trifluoroacetylpeptide anilide inhibitor. 888 99

The crystal structure of human neutrophil cathepsin G, complexed with the peptidyl phosphonate inhibitor Suc-Val-Pro-PheP-(OPh)2, has been determined to a resolution of 1.8 A using Patterson search techniques. The cathepsin G structure shows the polypeptide fold characteristic of trypsin-like serine proteinases and is especially similar to rat mast cell proteinase II. Unique to cathepsin G, however, is the presence of Glu226 (chymotrypsinogen numbering), which is situated at the bottom of the S1 specificity pocket, dividing it into two compartments. For this reason, the benzyl side chain of the inhibitor PheP residue does not fully occupy the pocket but is, instead, located at its entrance. Its positively charged equatorial edge is involved in a favourable electrostatic interaction with the negatively charged carboxylate group of Glu226. Arrangement of this Glu226 carboxylate would also allow accommodation of a Lys side chain in this S1 pocket, in agreement with the recently observed cathepsin G preference for Lys and Phe at P1. The cathepsin G complex with the covalently bound phosphonate inhibitor mimics a tetrahedral substrate intermediate. A comparison of the Arg surface distributions of cathepsin G, leukocyte elastase and rat mast cell protease II shows no simple common recognition pattern for a mannose-6-phosphate receptor-independent targeting mechanism for sorting of these granular proteinases.
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PMID:The 1.8 A crystal structure of human cathepsin G in complex with Suc-Val-Pro-PheP-(OPh)2: a Janus-faced proteinase with two opposite specificities. 889 42

The objective of this study was to characterize the plasmin-induced stimulation of leukotriene (LT) B4 biosynthesis in human peripheral monocytes (PM). Plasmin up to 175 x 10(-3) CTA U/ml triggers a concentration-dependent release of 5-lipoxygenase-derived LTB4 while release of the cyclooxygenase products thromboxane (TX) B2 and prostaglandin (PG) E2 remained unaffected. The stimulatory effect appeared to be specific in as much as 1) it was found in PM, but not in polymorphonuclear neutrophils (PMN), 2) it requires the lysine binding sites of plasmin molecule since it was inhibited by the lysine analogues 6-aminohexanoic acid (6-AHA) and trans-4(aminomethyl)cyclohexane-1-carboxylic acid (t-AMCA), 3) the intact catalytic center of plasmin is required since neither plasminogen nor catalytic center-blocked plasmin share the stimulatory effect of active plasmin, 4) other serine proteases such as alpha-chymotrypsin, human neutrophil elastase and cathepsin G did not stimulate release of detectable amounts of LTB4 from PM. In addition, catalytic center-blocked plasmin antagonized the stimulatory effect of active plasmin. Plasmin-mediated monocyte activation apparently proceeds via a pertussis toxin-sensitive G protein. Plasmin did not increase inositol (1,4,5) trisphosphate levels, but a time- and concentration-dependent stimulation of cyclic GMP formation was observed. The data show that plasmin is a specific stimulus for human peripheral monocytes. Plasmin may be an important link between the coagulation cascade and inflammatory reactions.
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PMID:Plasmin is a specific stimulus of the 5-lipoxygenase pathway of human peripheral monocytes. 890 97

The P1 or primary specificity residue of standard mechanism canonical protein inhibitors of serine proteinases, inserts into the S1 primary specificity cavity of the cognate enzyme upon enzyme-inhibitor complex formation. Both natural evolution and protein engineering often change the P1 residue to greatly alter the specificity and the binding strength. To systematize such results we have obtained all 20 coded P1 variants of one such inhibitor, turkey ovomucoid third domain, by recombinant DNA technology. The variants were extensively characterized. The association equilibrium constants were measured at pH 8.30, 21 (+/-2) degrees C, for interaction of these variants with six well characterized serine proteinases with hydrophobic S1, cavities. The enzyme names are followed by the best, worst and most specific coded residue for each. Bovine chymotrypsin A alpha (Tyr, Pro, Trp), porcine pancreatic elastase (Leu/Ala, Arg, Ala), subtilisin Carlsberg (Cys, Pro, Glu), Streptomyces griseus proteinase A (Cys, Pro, Leu) and B (Cys, Pro, Lys) and human leukocyte elastase (Ile, Asp, Ile). The data set was merged with Ka values for five non-coded variants at P1 of turkey ovomucoid third domain obtained in our laboratory by enzymatic semisynthesis. The ratios of the highest to the lowest Ka for each of the six enzymes range from 10(6) to 10(8). The dominant force for binding to these pockets is the hydrophobic interaction. Excess steric bulk (too large for the pocket), awkward shape (Pro, Val and Ile), polarity (Ser) oppose interaction. Ionic charges, especially negative charges on Glu- and Asp- are strongly unfavorable. The Pearson pro duct moment correlations for all the 15 enzyme pairs were calculated. We suggest that these may serve as a quantitative description of the specificity of the enzymes at P1. The sets of Streptomyces griseus proteinases A and B and of the two elastases are strongly positively correlated. Strikingly, chymotrypsin and pancreatic elastase are negatively correlated (-0.10). Such correlations can be usefully extended to many other enzymes and to many other binding pockets to provide a general measure of pocket binding specificity.
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PMID:Binding of amino acid side-chains to S1 cavities of serine proteinases. 904 74

Human myeloblastin (leukocyte proteinase 3) shares many biochemical properties with leukocyte elastase, but rapidly loses enzymatic activity when raising the pH and/or the ionic strength of an acidic solution or when handled in glass vessels. This poses limits to kinetic experiments requiring long incubation times. After purification, myeloblastin was conveniently stored in a glycine/HCl buffer at pH 3.2, while assays were performed in sodium/potassium phosphate buffer at pH 7.0, ionic strength 0.11, in the presence of 0.05% w/v Triton X-100 and taking care to avoid any contact with glass surfaces. The kinetic parameters of leukocyte elastase and myeloblastin with peptide substrates, irreversible inactivators and glycosaminoglycans were compared under these conditions. MeO-succinyl-Lys(2-picolinoyl)Ala-Pro-Val-4-nitroanilide, an excellent substrate for leukocyte elastase, also proved to be a good substrate for myeloblastin (Km = 16 microM, kcat/Km = 30,600 M(-1)s(-1)). Inactivation of myeloblastin by 3,4-dichloroisocoumarin (ki/Ki = 6,389 M(-1)s(-1)) and MeO-Suc-Ala-Ala-Pro-Val-chloromethane (ki/Ki = 579 M(-1) S(-1)) occurred via a two-step, irreversible complexing mechanism with potencies one-half and one-fifth that of leukocyte elastase, respectively. Glycosaminoglycans such as chondroitin sulfate, dermatan sulfate and a chondroitin polysulfate, interacted with myeloblastin as non-essential activators in the presence of peptide substrates (activation up to a 6.7-fold factor) and as partial inhibitors (about 50% inhibition at saturation) in the presence of elastin. This property distinguishes myeloblastin from leukocyte elastase, which is always inhibited by glycosaminoglycans, independently of the substrate.
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PMID:Human myeloblastin (leukocyte proteinase 3): reactions with substrates, inactivators and activators in comparison with leukocyte elastase. 906 56

A series of potent and selective human leukocyte elastase (HLE) inhibitors of the Val-Pro-Val type has been developed. Initially, the central proline residue was replaced by nonnatural amino acids Phi ((2S,3aS,7aS)-perhydroindole-2-carboxylic acid) and Abo ((3S)-2-azabicyclo-[2.2.2]octane-3-carboxylic acid), and secondly several groups able to confer antioxidant properties to the molecule were introduced at the lipophilic N-terminal side chain. When compared to reference inhibitors, in vitro HLE inhibitory potency was maintained (10-100 nM) both with compounds containing the antioxidant moiety at the end of the N-terminal side chain and with compounds in which the N-terminal valine of the tripeptidic sequence had been replaced by a epsilon-substituted lysine. The lipidic peroxidation inhibitory potency of this series of inhibitors was found to be similar to that of the reference antioxidant compounds (around 1 microM). Moreover, HLE-induced hemorrhage in the hamster lung was effectively prevented (40-60% at 15 micrograms/kg) by most of the inhibitors tested when administered intratracheally 3 h before instillation of elastase. Among the most active analogs, compounds 11a,c,g were still active when administered 18 h before elastase. Interestingly, compound 14a was able to prevent HLE-mediated lung damage when administered 72 h prior to enzymatic challenge, indicating exceptional stability and retention in the lung. Finally, in a 14-day chronic model of emphysema in the hamster, 14a significantly conserved alveolar spaces, a marker of lung tissue destruction, and was more potent than reference inhibitor ICI 200 880. This indicates that addition of peroxidation inhibitory properties to an HLE inhibitor can provide a powerful in vivo inhibitor of pulmonary tissue destruction.
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PMID:Dual inhibition of human leukocyte elastase and lipid peroxidation: in vitro and in vivo activities of azabicyclo[2.2.2]octane and perhydroindole derivatives. 919 69

The reactive center loop of native alpha1-proteinase inhibitor has been reported to be in a helical conformation and in a beta-strand conformation by two different studies. In the beta-strand loop structure the P5 glutamic acid plays a unique role by stabilizing the loop in the predicted optimal conformation for the interaction with target proteinases and insertion into beta-sheet A. We hypothesize here that disrupting the interactions that stabilize the beta-strand conformation of the loop would result in changes in the inhibitory properties of the serpin. In addition, our earlier studies on reactive center loop mutants of alpha1-proteinase inhibitor suggested that the P5 residue was important in stabilizing the alpha1-proteinase inhibitor-proteinase complexes. To address these issues we made mutants of alpha1-proteinase inhibitor with glycine, glutamine, or lysine at the P5 position and measured the rates and stoichiometries of inhibition with trypsin and human neutrophil elastase and the stabilities of the resulting complexes. In most cases the rate of inhibition was reduced by about half and the stoichiometry increased between 2- and 4-fold. The only exception was for trypsin with the lysine variant where the P5 was now the favored site of cleavage. These data show that the P5 Glu is important in maintaining the reactive center loop in a conformation optimal for interaction with the proteinase and for a fast rate of loop insertion. The complexes formed with trypsin and the variant serpins were less stable than that formed with wild-type serpin and resulted in up to 33% regeneration of trypsin activity over a period of 6 days, compared with 17% with wild type. Thus, the P5 residue of alpha1-proteinase inhibitor is important in all steps of the inhibitory mechanism in a manner consistent with the structural role played by this residue in the beta-strand loop structure of native alpha1-proteinase inhibitor.
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PMID:Influence of the P5 residue on alpha1-proteinase inhibitor mechanism. 946 13

Different families of protein inhibitors of serine proteases share similar conformation of the enzyme-binding loop, while their scaffoldings are completely different. In the enzyme-inhibitor complex, the P1position of the loop makes numerous contacts within the S1pocket and significantly influences the energy of the interaction. Here, we determine the association energies (DeltaGavalues) for the interaction of coded P1variants of bovine pancreatic trypsin inhibitor (BPTI) with bovine beta-trypsin (BT), anionic salmon trypsin (AST), bovine alpha-chymotrypsin (BCHYM), and human neutrophil elastase (HNE). The respective DeltaGaranges are 15, 13, 9, and 8 kcal mol-1(1 cal=4.18 J). Next, through interscaffolding additivity cycles, we compare our set of DeltaGavalues determined for BCHYM and HNE with similar data sets available in the literature for three other inhibitor families. The analysis of the cycles shows that 27 to 83 % of cycles fulfil the criteria of additvity. In one particular case (comparison of associations of P1variants of BPTI and OMTKY3 with BCHYM) there is a structural basis for strongly non-additive behaviour. We argue that the interscaffolding additvity depends on sequential and conformational similarities of sites where the mutation(s) are introduced and on the particular substitution. In the second interscaffolding analysis, we compare binding of the same P1mutants to BT and AST. The high correlation coefficient shows that both trypsins recognize with comparable strength the non-cognate side-chains. However, the cognate Arg and Lys side-chains are recognized significantly more strongly by AST.
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PMID:Interscaffolding additivity: binding of P1 variants of bovine pancreatic trypsin inhibitor to four serine proteases. 1033 15

The effect of modifications of Met, Arg, and Lys residues on the inhibitory activity of a serine proteinase-inhibiting 21-kD protein from potato tubers has been studied. The data indicate that the 21-kD protein has two independent reactive sites for human leukocyte elastase (or chymotrypsin) and trypsin. It is concluded that the 21-kD inhibitor has Met and Arg residues in the P1 position of the reactive sites responsible for interactions with elastase (or chymotrypsin) and trypsin. It is shown that the 21-kD protein is capable of forming a triple complex binding simultaneously one molecule of trypsin and one molecule of chymotrypsin.
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PMID:Reactive sites of the 21-kD protein inhibitor of serine proteinases from potato tubers. 1052 25

A 21-kD protein isolated earlier from potato tubers (Solanum tuberosum L.) has two isoforms, with pI 6.3 and 5.2, which were separated by fast protein ion-exchange chromatography on a Mono Q column. The primary structures of the two forms consisted of 187 and 186 amino acid residues. Both isoforms are composed of two polypeptide chains, designated A and B, linked by a single disulfide bond between Cys-146 of the A chain and Cys-7 of the B chain. The amino acid sequences of the A chains of the two forms, consisting of 150 residues each, differ in a single amino acid residue at position 52 (Val --> Ile), while the B chains, containing 37 and 36 residues, respectively, have substitutions at nine positions (Leu-8 --> Ser-8, Lys-25--Asp-26 --> Asn-25--Glu-26, Ile-31--Ser-32 --> Val-31--Leu-32, Lys-34--Gln-35--Val-36--Gln-37 --> Gln-34--Glu-35--Val-36). Both isoforms form stable inhibiting complexes with human leukocyte elastase and are less effective against chymotrypsin and trypsin.
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PMID:Primary structure of a 21-kD protein from potato tubers. 1061 30


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