共查询到20条相似文献,搜索用时 15 毫秒
1.
2.
M Vered Y Burstein A Gertler 《International journal of peptide and protein research》1985,25(1):76-84
Elastin was fully solubilized by digestion with elastase I or elastase II. Each digest was separated into high-molecular weight and low-molecular weight fractions that were characterized by the correspondence to their amino acid content, N-terminal sequence and C-terminal amino acids. It was found that although the relative amount of amino acids in the low-molecular weight fraction obtained by digestion with elastase I was lower than in digestion with elastase II, no major difference in the type of bonds cleaved in the low- or high-molecular weight fractions of each digest could be seen. There is, however, a remarkable difference in the type of bond cleaved by the two enzymes. While elastase I cleaves mostly Ala-Ala and also Ala-Gly bonds, elastase II hydrolyzes Leu-Ala, Leu-Gly, Phe-Ala, Phe-Gly and Tyr-Ala, Tyr-Gly bonds. Theoretical calculations led us to suggest both digests are composed of cross-linked peptides that vary not only in the molecular size but also in the number of cross-links found in peptides of the same size. 相似文献
3.
An electrophoretically homogeneous elastase preparation free from tryptic and chymotryptic activities was obtained by chromatography on DEAE-Sephadex and CM-cellulose. This preparation exhibits a narrower specificity towards peptide bonds than that observed by Naughton & Sanger (1961). With oxidized insulin B chain as substrate, the fastest breaks occur between alanine-14 and leucine-15 and between valine-18 and cysteic acid-19. The bond between glycine-23 and phenylalanine-24 is also efficiently hydrolysed. Other bonds hydrolysed are that between valine-12 and glutamic acid-13 and that between serine-9 and histidine-10. Oxidized insulin A chain is hydrolysed only at one of two points, between alanine-8 and serine-9 or between serine-12 and leucine-13, and the rate of hydrolysis is very low. 相似文献
4.
New evidence has enabled the identification of novel specific inhibitors of porcine pancreatic elastase with potential for action in vivo. The basis for optimization of activity in this series of derivatives of alanylproline, and the relationship of elastase inhibition to the binding mode, is discussed. 相似文献
5.
6.
7.
C Largman 《Biochemistry》1983,22(16):3763-3770
Proelastase has been purified to homogeneity from rat pancreatic tissue by a combination of CM-Sephadex and immobilized protease inhibitor affinity resins. Trypsin activation yields an elastolytic enzyme that possesses a specificity toward small hydrophobic residues in synthetic amide substrates, similar to those of porcine elastase 1 and canine elastase. However, the rat enzyme also rapidly hydrolyzes a substrate containing tyrosine in the P1 position. N-Terminal sequence analysis reveals that rat proelastase has an identical activation peptide with that of porcine proelastase 1 and has two conservative amino acid sequence differences from the activation peptide of canine proelastase. The sequence data established that rat proelastase corresponds to the elastase 1 mRNA clone isolated by MacDonald et al. [MacDonald, R. J., Swift, G. H., Quinto, C., Swain, W., Pictet, R. L., Nikovits, W., & Rutter, W. J. (1982) Biochemistry 21, 1453]. The sequence and substrate data obtained for rat and canine elastases suggest that there is a family of pancreatic elastases with properties similar to those of the classically described porcine elastase 1. 相似文献
8.
9.
Paolo Ascenzia Enea Menegatti Mario Guarneri Eraldo Antonini 《Molecular and cellular biochemistry》1983,56(1):33-38
Pre-steady-state and steady-state kinetics for the p.p. elastase-catalysed hydrolysis of ZAlaONp, one of the most favourable substrates for this serine protease, have been studied between pH 4.0 and 8.0. The results are consistent with the minimum three-step mechanism: (formula; see text) Under pre-steady-state conditions, where [E0] much greater than [S0], the values of the dissociation constant of the E X S complex (Ks = k-1/k+1) and of the individual rate constants for the catalytic steps (k+2 and k+3) have been determined over the whole pH range explored. Under steady-state conditions, where [S0] much greater than [E0], the values of kcat and Km have been obtained over the same pH range. The pH profiles of k+2, k+3, k+2/Ks, kcat, kcat/Km reflect the ionization of a group, probably His57, with a pKa value of 6.85 +/- 0.10. The values of Ks and Km are pH independent. The steady-state parameters for the p.p. elastase-catalysed hydrolysis of a number of p-nitrophenyl esters of N-alpha-carbobenzoxy-L-amino acids have been also determined between pH 4.0 and 8.0 and compared with those of b.beta-trypsin and b.alpha-chymotrypsin. For all the substrates examined the acylation step (k+2) is rate limiting in the p.p. elastase catalysis, between pH 4.0 and 8.0. The different catalytic behaviours of p.p. elastase, b.beta-trypsin and b.alpha-chymotrypsin are consistent with the known three-dimensional structures of these serine proteases. 相似文献
10.
M Scuri R Forteza I Lauredo J R Sabater Y Botvinnikova L Allegra W M Abraham 《Journal of applied physiology》2000,89(4):1397-1402
Neutrophil elastase has been linked to inflammatory lung diseases such as chronic obstructive pulmonary disease, adult respiratory distress syndrome, emphysema, and cystic fibrosis. In guinea pigs, aerosol challenge with human neutrophil elastase causes bronchoconstriction, but the mechanism by which this occurs is not completely understood. Our laboratory previously showed that human neutrophil elastase releases tissue kallikrein (TK) from cultured tracheal gland cells. TK has been identified as the major kininogenase of the airway and cleaves both high- and low-molecular weight kininogen to yield lysyl-bradykinin. Because inhaled bradykinin causes bronchoconstriction and airway hyperresponsiveness in asthmatic patients and allergic sheep, we hypothesized that elastase-induced bronchoconstriction could be mediated by bradykinin. To test this hypothesis, we measured lung resistance (RL) in sheep before and after inhalation of porcine pancreatic elastase (PPE) alone and after pretreatment with a bradykinin B(2) antagonist (NPC-567), the specific human elastase inhibitor ICI 200,355, the histamine H(1)-antagonist diphenhydramine hydrochloride, the cysteinyl leukotriene 1 receptor antagonist montelukast, or the cyclooxygenase inhibitor indomethacin. Inhaled PPE (125-1,000 microg) caused a dose-dependent increase in RL. Aerosol challenge with a single 500 microg dose of PPE increased RL by 132 +/- 8% over baseline. This response was blocked by pretreatment with NPC-567 and ICI-200,355 (n = 6; P < 0.001), whereas treatment with diphenhydramine hydrochloride, montelukast, or indomethacin failed to block the PPE-induced bronchoconstriction. Consistent with pharmacological data, TK activity in bronchial lavage fluid increased 134 +/- 57% over baseline (n = 5; P < 0.02). We conclude that, in sheep, PPE-induced bronchoconstriction is in part mediated by the generation of bradykinin. Our findings suggest that elastase-kinin interactions may contribute to changes in bronchial tone during inflammatory diseases of the airways. 相似文献
11.
Sakineh Sadeghi-Kaji Behzad Shareghi Ali Akbar Saboury Sadegh Farhadian 《Journal of biomolecular structure & dynamics》2020,38(1):78-88
AbstractThe aim of this study was to investigate the spermine effect on the thermal denaturation, conformation and activity of elastase at three temperatures of 303, 313 and 323?K in the Tris buffer, at pH 8.5, using UV–vis spectrophotometry, spectrofluorometry and circular dichroism as well as molecular docking and molecular simulation. The increased absorption of elastase in the presence of spermine suggested a change in the environment of tryptophan. It was found that under the influence of spermine, the emission intensity of elastase extremely was reduced, and the use of the Stern-Volmer equation showed that some static quenching had occurred. The thermodynamic parameters values (enthalpy and entropy) and the molecular docking technique also revealed that van der Waals forces or hydrogen bonding interactions played an important role in the binding process. The spermine–elastase complex formation led to increasing the value of the catalytic constant (kcat). So it could be considered as an activator. Slight changes were observed in the second structure of elastase (1.06% increase for the α-helix and 0.048% decrease the β-sheet) and the thermal stability effect. Molecular docking results also demonstrated that spermine could bind to porcine pancreatic elastase, and van der Waals forces or hydrogen bonding interactions played the major role in the binding process. Overall, our results showed that spermine could induce structural alterations in elastase, acting as a partial stabilizer and an activator for the enzyme.Communicated by Ramaswamy H. Sarma 相似文献
12.
W Ardelt 《Biochimica et biophysica acta》1975,393(2):267-273
Some molecular properties of the elastase II preparation, homogenous in ultracentrifugation, have been determined. The molecular weight is 25 000, the sedimentation coefficient and the diffusion coefficient are 3.69-10(-13) s(-1) and 12.09-10(-7) cm2/s, respectively. The partial specific volume was 0.716 g/cm3, and the axial ratio is 1.95. Elastase II exhibited a considerably lower content of arginine, tyrosine, and valine, and a higher content of proline, serine and conjugated carbohydrates than elastase I. The N-terminal amino acid of the enzyme is leucine, and its isoelectric point was 10.7. 相似文献
13.
The preparation and purification of tryptic peptides from aminoethylated Dip-elastase and [(14)C]carboxymethylated Dip-elastase, and of peptic peptides from native elastase is described. A summary of the results of chemical studies used to elucidate the amino acid sequence of these peptides is presented. Full details are given in a supplementary paper that has been deposited as Supplementary Publication SUP 50016 at the National Lending Library for Science and Technology, Boston Spa, Yorks. LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1973), 131, 1-20. These results, together with those from previously published papers, are used to establish the complete amino acid sequence of elastase, which is a single polypeptide chain of 240 residues, molecular weight 25900, containing four disulphide bridges. 相似文献
14.
We studied the proteolytic action in vitro of free and alpha 2-macroglobulin-bound porcine pancreatic elastase [EC 3.4.21.11] on the apolipoproteins of plasma: very low density lipoprotein (VLDL), low density lipoprotein (LDL), and high density lipoprotein (HDL). Polyacrylamide gel electrophoresis, isoelectric focusing and immunodiffusion tests of elastase-treated plasma lipoproteins revealed that apolipoprotein C-II and C-III polypeptides were more susceptible to elastase in free form than plasma apolipoproteins (A-I, A-II, B, and E). Elastase bound to alpha 2-macroglobulin did not show any such activities. 相似文献
15.
Y Shirasu H Yoshida T Mikayama S Matsuki J Tanaka H Ikenaga 《Journal of biochemistry》1986,99(6):1707-1712
We have cloned a DNA that is complementary to the messenger RNA that encodes porcine pancreatic elastase 1 from pancreas using rat pancreatic elastase 1 cDNA as a probe. This complementary DNA contains the entire protein coding region of 798 nucleotides which encodes an elastase of 266 amino acids, and 22 and 136 nucleotides of the 5' and 3'-untranslated sequences. When this deduced amino acid sequence was compared with known amino acid sequences, a carboxy-terminal 240 amino acids long peptide was found to be identical with a mature form of porcine pancreatic elastase 1, except for two amino acids. The porcine enzyme contains the same number of amino acid residues as the rat enzyme, and their amino acid sequences are 85% homologous. Taking the above findings together, we conclude that the cloned cDNA encodes a mature enzyme of 240 amino acids including a leader and activation peptide of 26 amino acids. We expressed the cloned porcine pancreatic elastase 1 cDNA in E. coli as a lac-fused protein. The resulting fused protein showed enzymatic activity and immunoreactivity toward anti-elastase serum. 相似文献
16.
L H Takahashi R Radhakrishnan R E Rosenfield E F Meyer D A Trainor M Stein 《Journal of molecular biology》1988,201(2):423-428
X-ray crystallographic data to 2.57 A resolution (1 A = 0.1 nm) have been measured for the complex of a peptidyl trifluoromethylketone inhibitor with porcine pancreatic elastase (PPE); R = 0.14. The inhibitor forms a stable complex with the enzyme by means of a covalent attachment to active site Ser195O gamma, resulting in a hemiketal moiety with tetrahedral geometry. The tripeptide protion binds as an antiparallel beta-sheet, with four hydrogen bonds augmenting the active-site covalent linkage, Ki = 9.5 microM. His57 exhibits a bifurcated H-bond to both Ser195O gamma and an F atom of the inhibitor. This study is one of a series which explores the binding geometry of a variety of small substrates and inhibitors to PPE. This peptidyl-PPE complex affords insight into the binding geometry of a novel trifluoromethylketone moiety to a serine proteinase. 相似文献
17.
Porcine pancreatic elastase (PPE) was crystallized in complex with a novel inhibitor at pH 5 and X-ray diffraction data were collected at a synchrotron source to 1.66 A. Crystals belong to the orthorhombic space group P2(1)2(1)2(1), with unit cell parameters a = 50.25 A, b = 57.94 A and c = 74.69 A. PPE is often used as model for drug target, due to its structural homology with the important therapeutic target human leukocyte elastase (HLE). Elastase is a serine protease that belongs to the chymotrypsin family, which has the ability to degrade elastin, an important component in connective tissues. Excessive elastin proteolysis leads to a number of pathological diseases. 相似文献
18.
The primary structure of porcine pancreatic elastase: The N-terminus and disulphide bridges 总被引:4,自引:4,他引:0
下载免费PDF全文
![点击此处可从《The Biochemical journal》网站下载免费的PDF全文](/ch/ext_images/free.gif)
1. The amino acid composition and N-terminal groups of purified elastase show that it is a single peptide chain of 234 residues. 2. The N-terminal sequence is Val-Val-Gly-Gly-Thr-Glu-. 3. The sequences around the four disulphide bridges were determined by using a ;diagonal' electrophoretic technique. 4. These four bridges are homologous with the four common to bovine trypsin and chymotrypsin. 5. Out of 83 residues of the elastase sequence so far determined, 43 are homologous with similar regions of trypsin and chymotrypsin. 6. The evolutionary ancestry of these enzymes is discussed. 相似文献
19.
20.
Prior to availability of the crystal structure of the complex, we evaluated models of the complex between porcine pancreatic elastase and a t-Boc–Val-derived benzoxazinone inhibitor. Models of the noncovalent and covalent complex were generated using computer graphics and each model was subjected to energy minimization using molecular mechanics. After the crystal structure became available, we found that the model with the lowest energy was in good agreement with the crystal structure, except for the position of the His57 side chain. Permissible conformations of the inhibitor were based on information from x-ray crystal structures and an earlier conformational energy investigation of t-Boc–amino acids. We did not, however, limit ourselves to these conformations. The conformation of the inhibitor in the lowest energy model and crystal structure, was not similar to any of the minimum-energy conformations of t-Boc–amino acids. This suggests that limiting proposed binding modes only to the lowest energy conformations of a ligand (prior to binding) may sometimes unfairly bias the procedure. 相似文献