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)

In solution, MeO-Suc-Ala-Ala-Pro-D,L-boro-Val pinacol ester (Boroval) is a highly effective but reversible inhibitor of both porcine pancreatic elastase and human neutrophil elastase (HNE) (50% inhibition with a 1.5 M ratio of Boroval to elastase). Boroval has been shown to prevent porcine-pancreatic-elastase-induced emphysema in hamsters. But with HNE-induced emphysema in hamsters, pretreatment with as much as a 170-fold M excess of Boroval, given intratracheally 1 h before 0.3 mg HNE, did not prevent emphysema. Indeed, lung volumes were larger after Boroval pretreatment than after HNE alone. Emphysema was also induced by instilling HNE that had been mixed with and inactivated by a 41-fold M excess of Boroval (a molar ratio of 42). When 0.25 or 0.5 mg of HNE were given mixed with a 41-fold M excess of Boroval, the emphysema was much more severe with the 0.5 mg dose. Two hours after instillation of 0.3 mg HNE inactivated with a 34-fold M excess of Boroval, bronchoalveolar lavage contained elastolytic activity but no evidence of hemorrhage. In contrast, hemorrhage was severe in hamsters that had been instilled with 0.3 mg HNE alone. We conclude that Boroval can enhance HNE-induced emphysema. We postulate that Boroval suppresses HNE-induced hemorrhage and the resultant influx of plasma protease inhibitors; the HNE-Boroval complex is transported into the alveolar interstitium, followed by dissociation of the inhibitor from the active site of HNE. Because of its small size, free Boroval is rapidly cleared, and the reactivated HNE attacks elastic fibers, giving rise to emphysema.
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PMID:Induction and exacerbation of emphysema in hamsters with human neutrophil elastase inactivated reversibly by a peptide boronic acid. 229 86

alpha 1-Antitrypsin (alpha 1AT), the major serum inhibitor of neutrophil elastase, is a highly polymorphic protein associated with isoelectric focusing (IEF) patterns typical for each variant. alpha 1AT Vmunich, a previously unreported normal alpha 1AT variant, has a unique IEF banding pattern in which the 7 and 8 alpha 1AT protein bands focus with the normal M-type 7 and 8 bands, despite the fact that the major fraction of the Vmunich protein focuses in the "V" region of the IEF gel. To characterize the molecular basis of this variant and its unique IEF pattern, DNA sequence analysis of the coding exons of the Vmunich alpha 1AT gene was carried out using the polymerase chain reaction. The Vmunich allele differed from the common normal M1(Val213) alpha 1AT allele by a single nucleotide substitution of cytosine for adenosine, with the resultant amino acid change Asp2 GAT----Ala GCT. Inheritance of the allele was confirmed by family analysis using allele-specific amplification with the polymerase chain reaction. The Asp2----Ala mutation explains the cathodal position of the Vmunich protein on IEF, as there is a substitution of a negatively charged amino acid by a neutral one.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Characterization of the normal alpha 1-antitrypsin allele Vmunich: a variant associated with a unique protein isoelectric focusing pattern. 231 26

We have synthesized a series of peptidyl fluoroketones that reversibly inhibit the serine proteases human leukocyte elastase (HLE) and alpha-lytic protease (alpha-LP). Ac-ambo-AlaCF3 (1) inhibits HLE and alpha-LP with Ki's of 2.4 and 15 mM, respectively. The effects of structural variations on this parent compound on Ki and the kinetics of inhibition were studied. The acetyl group was replaced by the tripeptide Z-L-Ala-L-Ala-L-Pro to yield the tetrapeptide trifluoroketone (TFK) Z-L-Ala-L-Ala-L-Pro-ambo-AlaCF3 (2). This extension reduced Ki 3500-fold for HLE and 3000-fold for alpha-LP. Removal of a fluorine atom from a TFK decreases Ki about 15- to 30-fold with both enzymes. Replacement of one fluorine atom of 2 by a residue (-CH2-CH2-COLeuOMe) (6) which can interact with the S'1 and S'2 subsites decreased Ki 30-fold for HLE and 150-fold for alpha-LP compared to Z-L-Ala-L-Ala-L-Pro-ambo-AlaCF2H (3). The Ki of 6 for HLE is approximately equal to that of trifluoroketone 2. For alpha-LP Ki of 6 is 10-fold lower than that for the trifluoroketone 2. Inhibitors with Ki values less than 10(-7) M exhibit slow binding kinetics. By analogy to cholinesterases and chymotrypsin, it is likely that these enzymes combine with the keto form of the inhibitor to form the enzyme-inhibitor complex. Therefore, kon and Ki were corrected for the ketone concentration. The corrected kon values for the slow binding inhibitors are in most cases less than diffusion controlled, ranging between 8.2 X 10(4) and 4.68 X 10(6) M-1 s-1. An exception is Z-L-Ala-L-Ala-L-Pro-ambo-ValCF3 (8) where kon = 9 X 10(7) M-1 s-1, which is nearly diffusion controlled.
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PMID:Structure-activity studies of fluoroketone inhibitors of alpha-lytic protease and human leukocyte elastase. 235 15

The semisynthesis of homologues of aprotinin (BPTI) is described. The P1 amino acid residue of these homologues was substituted by other amino acids using peptide synthetic methods. The reactive-site-modified inhibitor (with the Lys15-Ala16 peptide bond hydrolyzed) was used as starting material. All carboxyl groups of the modified inhibitor were esterified with methanol, then the Lys15 methyl ester group was hydrolyzed selectively. Afterwards, Lys15 itself was split off. A new amino acid residue was incorporated by using water-soluble carbodiimide combined with an acylation catalyst. tert-Butyl-ester-protected amino acids were used for reinsertion. The method was tested by re-insertion of Lys15 to reconstitute the original inhibitor. Thirteen BPTI homologues with coded (Lys, Glu, Gly, Ala, Val, Ile, Leu) or uncoded amino acids (Abu, Ape, aIle, Ahx, tLeu, Neo) in position 15 were synthesized and the specificity of the inhibitors investigated. Amongst these, [Val15]BPTI was shown to be an excellent inhibitor for human polymorphonuclear leukocyte elastase having a complex dissociation constant of 0.11 nM. This inhibitor showed no detectable affinity to bovine pancreatic trypsin.
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PMID:Preparation of chemically 'mutated' aprotinin homologues by semisynthesis. P1 substitutions change inhibitory specificity. 245 25

The replacement of ser359 with ala359 at the P1 position in human alpha-1-proteinase inhibitor results in the production of a variant protein containing 15% of the inhibitory activity of the normal inhibitor. Separation of active from inactive inhibitor on anhydrochymotrypsin-sepharose yields a form which has a second order association rate with neutrophil elastase which is approximately one half that for the native protein. These data indicate that the P1 residue is not of primary importance during the interaction of proteinases with alpha-1-proteinase inhibitor. Since substitution of alanine for serine causes the formation, primarily, of inactive inhibitor the major function of ser359 probably involves proper folding to give a functionally active inhibitory conformation.
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PMID:The primary role of the P1 residue (ser359) of alpha-1-proteinase inhibitor. 249 88

Pseudomonas aeruginosa, which may cause severe lung infections, secretes a metalloelastase that may interfere with the assay of neutrophil elastase and cathepsin G in lung secretions. Using nuclear magnetic resonance spectroscopy, we have shown that P. aeruginosa elastase (PsE) cleaves succinyl-Ala3-p-nitroanilide between the first and the second alanine residue, rendering this substrate inefficient for the assay of neutrophil elastase. The cleavage occurs with a kcat/Km of 2.4 X 10(3) M-1 s-1, a value eightfold higher than the kcat/Km for the chromogenic cleavage of succinyl-Ala3-p-nitroanilide by neutrophil elastase. P. aeruginosa elastase also cleaves the elastase substrate succinyl-Ala3-Val-p-nitroanilide between the second and the third alanine residue and the cathepsin G substrate succinyl-Ala2-Pro-Phe-p-nitroanilide at the Pro-Phe linkage. By contrast, methoxysuccinyl-Ala2-Pro-Val-p-nitroanilide, another elastase substrate, is not hydrolyzed by the bacterial enzyme. Our data indicate that synthetic substrates should be used with caution to assay elastase and cathepsin G in lung secretions or other biologic fluids in which metalloproteinases may be present.
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PMID:Nonchromogenic hydrolysis of elastase and cathepsin G p-nitroanilide substrates by Pseudomonas aeruginosa elastase. 251 50

Nonporous, microparticulate, monodisperse silicas with particle diameters between 0.7 and 2.1 microns are introduced as stationary phases in high-performance affinity chromatography. The immobilization of m-aminophenylboronic acid, p-aminobenzamidine, tri-L-alanine, and concanavalin A onto these silicas was successfully achieved using 3-isothiocyanatopropyl-triethoxysilane as an activation reagent. Immobilized phenylboronic acid was applied to the isolation of nucleosides, nucleotides, and glycoprotein hormones such as bovine follicotropin and human chorionic gonadotropin, while immobilized benzamidine was employed for the isolation of the serine proteases thrombin and trypsin, immobilized tri-L-alanine for the separation of pig pancreatic elastase and human leukocyte elastase, and immobilized concanavalin A for the isolation of horseradish peroxidase. In all affinity chromatographic systems studied, the nonporous monodisperse silicas showed improved chromatographic performance compared to results obtained with porous silica supports using identical activation and immobilization procedures. Furthermore, frontal analysis was used as a method to evaluate the influence of experimental parameters on biological activity and accessible ligand densities. Only minor changes in bioactivity were found with the nonporous affinity supports, where accessibilities were typically higher than ca. 60%. The immobilization of affinity ligands onto porous supports as used in this and associated papers thus represents a successful general procedure for the preparation of stable matrices with fast kinetics for use in high-performance affinity chromatography.
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PMID:High-performance liquid affinity chromatography with phenylboronic acid, benzamidine, tri-L-alanine, and concanavalin A immobilized on 3-isothiocyanatopropyltriethoxysilane-activated nonporous monodisperse silicas. 254 22

alpha 1-Antitrypsin (alpha 1AT) is a highly pleomorphic 52-kDa serum glycoprotein that functions as the major inhibitor of neutrophil elastase. Of these, the most common normal alpha 1AT haplotypes identified by isoelectric focusing (IEF) of serum are those of the M family, including M1, M2, and M3. In the course of studying the alpha 1AT type Z gene, we identified a restriction endonuclease BstEII polymorphism in the M1 gene that predicted the existence of a previously unidentified, but relatively common, haplotype of M, referred to as M1(Ala213) [Nukiwa, T., Satoh, K., Brantly, M. L., Ogushi, F., Fells, G. A., Courtney, M., & Crystal, R. G. (1986) J. Biol. Chem. 261, 15989-15994]. In this study we have cloned both alpha 1AT genes from an individual heterozygous for the M1(Ala213) and M1(Val213) haplotypes. Sequencing of the coding exons of both demonstrated that they are identical except for the Ala-Val difference at residue 213. The codominant transmission of the M1(Ala213) gene was demonstrated in a family study. Evaluation of 39 genomic samples of Caucasians with the IEF haplotype M1 demonstrated haplotype frequencies of 68% for M1(Val213) and 32% for M1(Ala213). alpha 1AT serum levels of individuals inheriting the M1(Ala213) gene in a homozygous fashion were in the same range as those for homozygous M1(Val213) as was the rate of association of the M1(Ala213) protein with neutrophil elastase.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Characterization of the M1(Ala213) type of alpha 1-antitrypsin, a newly recognized, common "normal" alpha 1-antitrypsin haplotype. 289 Mar 73

Human neutrophil elastase (HNE) has been implicated as a major contributor to tissue destruction in various disease states, including emphysema. The structure of HNE, at neutral pH, in complex with methoxysuccinyl-Ala-Ala-Pro-Ala chloromethyl ketone (MSACK), has been solved and refined to an R factor of 16.4% at 1.84-A resolution. Results are consistent with the currently accepted mechanism of peptide chloromethyl ketone inhibition of serine proteases, in that MSACK cross-links the catalytic residues His-57 and Ser-195. The structure of the HNE-MSACK complex is compared with that of porcine pancreatic elastase in complex with L-647,957, a beta-lactam inhibitor of both elastases. The distribution of positively charged residues on HNE is highly asymmetric and may play a role in its specific association with the underlying negatively charged proteoglycan matrix of the neutrophil granules in which the enzyme is stored.
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PMID:Structure of human neutrophil elastase in complex with a peptide chloromethyl ketone inhibitor at 1.84-A resolution. 291 84

The enzyme elastase (EC 3.4.21.37) has proved to be a convenient and extremely sensitive marker for the quantification of neutrophils in cutaneous infiltrates. Fluorometric assay using the synthetic substrate MeOSuc-Ala-Ala-Pro-Val-N-methylcoumarin permitted the measurement of this enzyme in as few as five cells and was linear up to about 1000 cells per sample. The mean activity of lysates of human blood-derived neutrophils was 0.57 +/- 0.08 pmol of 7-amino-4-methyl-coumarin released per hour per neutrophil. Extracts of normal human skin contained no measurable elastase activity but resulted in a slight inhibition of the neutrophil enzyme (mean 12%). Application to the in vivo situation has been demonstrated by the use of leukotriene B4 as chemotactic agent. A reproducible neutrophil infiltrate was found at a dose of 2 ng, well below the threshold for the appearance of microabscesses.
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PMID:Elastase, a marker for neutrophils in skin infiltrates. 301


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