Elastases specificity

Elafin (12), an elastase-specific inhibitor isolated from human skin, is a 57-residue peptide containing four intramolecular disulfide bonds (Scheme 7).181,821... 

O. Wieddow, J. M. Schroder, H. Gregory, J. A. Young, and E. Christophers. Elafin an elastase-specific inhibitor of human skin. J. Biol. Chem. 265 14791(1990). 

Q.-L. Ying and S. R. Simon. Kinetics of the inhibition of human leukocyte elastase by elafin, a 6-kilodalton elastase-specific inhibitor from human skin. Biochemistry 32 1866(1993). 

In the recent studies, the enzyme shows that the overall polypeptide fold of chymotrypsin-like serine protease possesses essential SI specificity determinants characteristic of elastase using the multiple isomorphous replacement (MIR) method and refined to 2.3 A resolution Fig. (5). Structure-based inhibitor modeling demonstrated that EFEa s SI specificity pocket is preferable for elastase-specific small hydrophobic PI residues, while its accommodation of long and/or bulky PI residues is also feasible if enhanced binding of the substrate and induced fit of the SI pocket are achieved [Fig. (6) shows the active sites of serine protease]. EFEa is thereby endowed with relatively broad substrate specificity, including the dual fibrinolysis. This structure is the first report of an earthworm fibrinolytic enzyme component, a serine protease originated from annelid worm [17]. 

Elastase is a serine protease that is a protein target for drugs used in the treatment of emphysema. Elastase is so named because it rapidly hydrolyzes the otherwise nearly indigestible protein elastin, which is rich in Ala, Gly, and VaL Elastase specifically cleaves peptide bonds following small nonpolar residues, particularly Ala. 

Sallenave JM, Xing Z, Simpson AJ, Graham FL, Gauldie J. Adeno virus-mediated expression of an elastase-specific inhibitor (elafin) a comparison of different promoters. Gene Ther 1998 5 352-360. 

This is nicely illustrated by members of the chymotrypsin superfamily the enzymes chymotrypsin, trypsin, and elastase have very similar three-dimensional structures but different specificity. They preferentially cleave adjacent to bulky aromatic side chains, positively charged side chains, and small uncharged side chains, respectively. Three residues, numbers 189, 216, and 226, are responsible for these preferences (Figure 11.11). Residues 216... 

Figure 11.11 Schematic diagrams of the specificity pockets of chymotrypsin, trypsin and elastase, illustrating the preference for a side chain adjacent to the scisslle bond In polypeptide substrates. Chymotrypsin prefers aromatic side chains and trypsin prefers positively charged side chains that can interact with Asp 189 at the bottom of the specificity pocket. The pocket is blocked in elastase, which therefore prefers small uncharged side chains.

Trypsin, chymotrypsin, and elastase all carry out the same reaction—the cleavage of a peptide chain—and although their structures and mechanisms are quite similar, they display very different specificities. Trypsin cleaves peptides... 

In the past number of years a number of studies have shown that in a variety of diseases there is a significant oxidation of Met residues to Met(O) in specific proteins that results in a loss of biological activity. These diseases include cataracts, rheumatoid arthritis, adult respiratory distress syndrome and emphysema. The most convincing evidence that Met(O) in proteins may be involved in the etiology of a pathological condition comes from studies with a-l-PI. It is well accepted that a-l-PI is inactivated upon oxidation of its Met residues. A decreased activity of a-l-PI in lung tissue that would result in an increased elastase activity has been associated with pulmonary emphysema. In patients who have a... 

Transition state theory, 46,208 Transmission factor, 42,44-46,45 Triosephosphate isomerase, 210 Trypsin, 170. See also Trypsin enzyme family active site of, 181 activity of, steric effects on, 210 potential surfaces for, 180 Ser 195-His 57 proton transfer in, 146, 147 specificity of, 171 transition state of, 226 Trypsin enzyme family, catalysis of amide hydrolysis, 170-171. See also Chymotrypsin Elastase Thrombin Trypsin Plasmin Tryptophan, structure of, 110... 

There are two main classes of proteolytic digestive enzymes (proteases), with different specificities for the amino acids forming the peptide bond to be hydrolyzed. Endopeptidases hydrolyze peptide bonds between specific amino acids throughout the molecule. They are the first enzymes to act, yielding a larger number of smaller fragments, eg, pepsin in the gastric juice and trypsin, chymotrypsin, and elastase secreted into the small intestine by the pancreas. Exopeptidases catalyze the hydrolysis of peptide bonds, one at a time, fi"om the ends of polypeptides. Carboxypeptidases, secreted in the pancreatic juice, release amino acids from rhe free carboxyl terminal, and aminopeptidases, secreted by the intestinal mucosal cells, release amino acids from the amino terminal. Dipeptides, which are not substrates for exopeptidases, are hydrolyzed in the brush border of intestinal mucosal cells by dipeptidases. 

The proteases are secreted as inactive zymogens the active site of the enzyme is masked by a small region of its peptide chain, which is removed by hydrolysis of a specific peptide bond. Pepsinogen is activated to pepsin by gastric acid and by activated pepsin (autocatalysis). In the small intestine, trypsinogen, the precursor of trypsin, is activated by enteropeptidase, which is secreted by the duodenal epithelial cells trypsin can then activate chymotrypsinogen to chymotrypsin, proelas-tase to elastase, procarboxypeptidase to carboxypepti-dase, and proaminopeptidase to aminopeptidase. 

Serine proteases usually show primary specificity (occupation of subsite Si) for positively charged arginine or lysine (trypsin, plasmin, plasminogen activators, thrombin), large hydrophobic side chains of phenylalanine, tyrosine, and tryptophan (chymotrypsin, cathepsin G, chymase, and subtilisin), or small aliphatic side chains (elastases). Nevertheless, there are a large number of variations and in many cases, other subsites like S2 and S3 are more discriminating while maintaining the... 

Finally, coumarin derivatives may act as general inhibitors of serine proteases or as specific inhibitors of human leukocyte elastase, depending on the nature of the substituents, through two distinct mechanisms, suicide substrates (a-chymotrypsin)... 

Doucet, C. Pochet, L. Thierry, N. Pirotte, B. Delarge, J. Reboud-Ravaux, M. 6-Substituted 2-OXO-2H-1 -benzopyran-3-carboxyI ic acid as a core structure for specific inhibitors of human leukocyte elastase. J. Med. Chem. 1999, 42, 4161—4171. 

IL-lra (17.5) Monocyte/macrophage, fibroblast Specifically inhibits IL-1 effects, including SIRS and sepsis in animal models and humans. Attenuation of coagulation, fibrinolytic, and complement systems, levels of PAF and neutrophil elastase. 

Pohl, J., Pereira, A., Martin, N. M., Spitznagel, J. K. (1990). Amino acid sequence of CAP37, a human neutrophil granule-derived antibacterial and monocyte-specific chemotactic glycoprotein structurally similar to neutrophil elastase. FEBS Lett. 272, 200-4. 

C5a is inactivated by the myeloperoxidase-H202 system, which oxidises a methionine residue (Met 70) on the molecule group A streptococcal endo-proteinases also abolish chemotactic activity of C5a and related compounds. Neutrophil lysosomal enzymes (e.g. elastase and cathepsin G) also destroy C5a chemotactic activity, but as these proteases are inhibited by the serum antiproteinases, a -antiproteinase and a2-macroglobulin, the physiological role of neutrophilic proteases in the inactivation of C5a is questionable. Two chemotactic factor inactivators have been found in human serum an a-globulin that specifically and irreversibly inactivates C5-derived chemotactic factors, and a / -globulin that inactivates bacterial chemotactic factors. These activities are heat labile (destroyed by treatment at 56 °C for 30 min) and are distinct from those attributable to anaphylatoxin inactivator. An apparently specific inhibitor of C5-derived chemotactic activity has also been described in human synovial fluid and peritoneal fluid. This factor (molecular mass of 40 kDa) is heat stable and acts directly on C5a. 

N-Carbobenzoxy-L-alanine-/>-nitrophenyl ester is a specific substrate for elastase in which the rate-limiting step is deacylation, that is, hydrolysis of the acyl-enzyme intermediate. In 70% methanol over a reasonable temperature range the energy of activation of the turnover reaction, that is, deacylation, is 15.4 kcal mol. In the pH 6-7 region in this cryoprotective solvent, the turnover reacdon can be made negligibly slow at temperatures of -50 C or below. Under such conditions/i-nitro-phenol is released concurrent to acyl enzyme formation in a 1 1 stoichiometry with active enzyme in the presence of excess substrate. In other words, even at low temperatures, the acylation rate is much faster than deacylation and the acyl enzyme will accumulate on the enzyme. The rate of acyl-enzyme formation can be monitored by following the rate of p-nitrophenol release, and thus the concentration of trapped acyl enzyme may be determined. This calculadon has been carried out and... 

Trypsin, chymotrypsin, and elastase are en-dopeptidases that belong to the group of serine proteinases (see p. 176). Trypsin hydrolyzes specific peptide bonds on the C side of the basic amino acids Arg and Lys, while chymotrypsin prefers peptide bonds of the apolar amino acids Tyr, Trp, Phe, and Leu (see p. 94). 

Adjacent half-cystine residues are present in many peptides and proteins. In most of the cases they form two disulfide bonds with other cysteines in the molecule, unless the peptide bond between them is cis (see Section 6.1.5.1 ).t40 41 Specific enzymes that cleave the Cys-Xaa bond have not yet been discovered, although there are a few reports of cleavage of the Cys-Cys bond by enzymes such as elastase and pepsinJ42-43 For peptides with Cys-Cys bonds the cleavage method in Section 6.1.6.2.4 is recommended. 

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