Porcine pancreatic elastase

Human leukocyte elastase is a protease that degrades elastin and other connective tissue components. It is implicated in the pathogenesis of pulmonary emphysema and other inflammatory diseases such as rheumatoid arthritis and cystic fibrosis. Porcine pancreatic elastase has often been used as a model for HLE. Both enzymes have a small primary binding site Si. 

Vijayalakshmi, J. Meyer, E. F. Kam, C.-M. Powers, J. C. Structural study of porcine pancreatic elastase complexed with 7-amino-3-(2-bromoethoxy)-4-chloroisocoumarin as a nonreactivable doubly covalent enzyme-inhibitor complex. Biochemistry 1991, 30, 2175-2183. 

The ultimate objective of an X-ray cryoenzymological study is the mapping of the structures of all kinetically significant species along the reaction pathway. In the case of ribonuclease A this has been largely achieved, as described above. Other enzymatic reactions now await application of the same techniques. Unfortunately, not all crystalline enzymes lend themselves to study by this method. In some cases it may be impossible to find a suitable cryoprotective mother liquor in others, the reaction may occur too rapidly at ordinary temperature. A reaction with Acat of 10 seconds and an activation enthalpy of —6 kcal mol will not be quenched even at — 75°C. The approach we have described in this article can be applied to only a small number of enzymes. Two likely candidates for successors to ribonuclease are the enzymes yeast triosephosphate isomerase and porcine pancreatic elastase. 

L.H. Takashashi, R. Radhakrishnan, R.E. Rosenfield, E.F. Meyer, D.A. Trainor, Crystal structure of the covalent complex formed by a peptidyl a,a-difluoro- -keto amide with porcine pancreatic elastase at 1.78 A resolution, J. Am. Chem. Soc. Ill (1989) 3368-3374. 

Klee (74) has shown that porcine pancreatic elastase has an effect on RNase-A at pH 8 similar to subtilisin. In this case Ala 20 is excised by... 

Powers, J. C., et al. 1977. Specificity of porcine pancreatic elastase, human leukocyte elastase and cathepsin G. Inhibition with peptide chloromethyl ketones. Biochim Biophys Acta 485 156. 

The electrospray ionization mass spectrometry (ESI-MS) analysis of the incubation between porcine pancreatic elastase (PPE) and the tert-butylammonium salt of clavulanic acid 54 (R1 = R2 = H) at time points of between 3 min and 5h revealed that there were no mass increments relative to PPE. However, when the benzyl derivative 54 (R1 = Bn, R2 = H) was used, a peak was observed at 26187 Da after 3 min, which corresponded to the formation of an initial acyl-enzyme complex. The intensity of the peak decreased significantly and two clear additional peaks at 25 968 and 25 967 Da appeared after 5 min, which corresponded to adducts with mass increments at 77 and 88 Da, respectively. The intensities of both peaks decreased after 60 min and almost disappeared after 5 h. The corresponding />-nitrobenzyl ester 54 (R1 = CH2PhN02, Rz = H) showed similar results except that the formation of adducts appeared slightly faster <2000T5729>. 

Human aranti-trypsin (AAT) 22 mgh1 Inhibition of porcine pancreatic elastase Heterogeneous [17]... 

Powers JC, Oleksyszyn J, Narasimhan SL, Kam C-M, Radhakrishnan REF, Meyer J (1990) Reaction of Porcine pancreatic elastase with 7-substituted 3-alkoxy-4-chloroisocoumarins design of potent inhibitors using the crystal structure of the complex formed with 4-chloro-3-ethoxy-7-guanidinoisocoumarin. Biochemistry 29 3108-3118... 

Sawyer L, Shotton DM, Campbell JW, Wendell PL, Muirhead H, Watson HC, Diamond R, Ladner RC (1978) The atomic structure of crystalline porcine pancreatic elastase at 2.5 A resolution Comparison with the structure of a-chymotrypsin. J Mol Biol 118 137-208... 

Understanding how HLE inhibitors work and/or designing new inhibitors requires a model of HLE s active-site and an understanding of its mechanism of action. All serine proteinases share a similar catalytic region and mechanism of action but differ in several amino acids in the extended substrate-binding region. These changes are responsible for the specificity differences between HLE and other serine proteinases. In some cases analysis of the enzyme-inhibitor interactions has only been carried out with other related enzymes, and those results are referenced as appropriate. One closely related enzyme, porcine pancreatic elastase (PPE, EC 3.4.21.36) has... 

Abbreviations used in tables include Ada = 1-adamantyl boro-AA-OH = boronic acid analogue of specified amino acid (aa) Cbz = benzyloxycarbonyl CMK = chloromethylketone EIM = enzyme inhibitor of monocytes -NA = p-nitroanilide ND = not determined NR = not reactive MeO-Suc = methoxy succinoyl Met(O) = methionine sulphoxide Pic = picolinyl PPE = porcine pancreatic elastase SLPI = secretory leukocyte proteinase inhibitor TFMK = trifluoromethylketone Z = benzyloxycarbonyl. 

Scyptolins A and B (Fig. 48) were isolated from axenic cultures of the terrestrial cyanobacterium Scytonema hofmanni PCC7110 by Matem et al. in 2001.They have the uncommon amino acid residue 3 -chloro-jV-methyl tyrosine and unique side chains. Both scyrtolin A and B exhibit selective inhibition of porcine pancreatic elastase in vitro with an IC50 of 3.1pg/ml l... 

Figure 5. Variation of the protein MEP along the active sites of some enzymes. It a-chymotiypsin, 2t p-tiypsin, 3 porcine pancreatic elastase, 4 Streptomyces Griseus hydrolase, Si a-lytic protease, 6t subtilisin NOVO, 7i acetylcholinesterase, 8> lipase A, 9 lysozyme, lOi D-xyloie isomerase. Point A is at OG of die active serine in 1-8, at the bisector ofODl and OD2 of Asp-52 in 9, at Ol of the cyclic xylose m 10. Point B is atNE2 of the catalytic histidine in 1-8, in the first trisector of points A and Din 9, atNEl ofHis-S4in 10. Point C is at ND1 of the catalytic histidine in 1-8 and 10, at the second trisector of points A and D in 9. Point D is at the bisector of the carboxylate oxygens of the catalytic Asp or Glu side chains.

Mattos, C., Giammona, D. A., Petsko, G. A. and Ringe, D. (1995) Structural analysis of the active site of porcine pancreatic elastase based on the X-ray crystal structures of complexes with trifluoroacetyl-dipeptide-anilide inhibitors. Biochemistry, 34, 3193-3203. 

Peet, N. P, Burkhart, J. P., Angelastro, M. R., et al. (1990) Synthesis of peptidyl fluoromethyl ketones and peptidyl a-keto esters as inhibitors of porcine pancreatic elastase, human neurophil elastase, and rat and human neurophil cathepsin. J. Med. Chem., 33, 394-407. 

Some protoberberine and structurally related alkaloids were tested for inhibitory activity on porcine pancreatic elastase (PPE) and human sputum elastase (HSE). Berberine chloride significantly inhibited the elastolytic activity of both enzymes, but tetrahydroberberine had no effect. It appears that the quaternary nitrogen atom of these alkaloids plays an important role in the inhibition of elastolytic activity. The amidolytic activity of the elastases was not affected by any of the test alkaloids [240]. 

In a similar case, Navia et al. (261) solved the structure of the complex of the irreversible inhibitor 33 with porcine pancreatic elastase at 1.84 A resolution. 

Pharmaceutical companies are increasingly interested in developing products based on proteins, enzymes, and peptides. With the development of such products comes the need for methods to evaluate the purity and structural nature of these biopharmaceuticals. Proteins, unlike traditional pharmaceutical entities, rely on a specific secondary structure for efficacy. Methods to monitor the secondary structure of pharmaceutically active proteins, thus, is necessary. Infrared spectroscopy provides a way to study these compounds quickly and easily. Byler et al. (65) used second-derivative IR to assess the purity and structural integrity of porcine pancreatic elastase. Seven different lyophilized samples of porcine pancreatic elastase were dissolved in D20, placed in demountable cells with CaF2 windows, and IR spectra obtained. The second derivatives of the spectra were calculated and the spectral features due to residual water vapor and D20 removed. 

Lyophilized porcine pancreatic elastase exhibited denaturation during storage at 40°C and 75% relative humidity in which oxidation of tryptophyl groups was probably involved.823 Reducing sugar impurities in mannitol used as an excipient824 induced solid-state oxidative degradation of a cyclic heptapeptide. 

Porcine pancreatic elastase-t-peptidyl boronic acid FAST 51.78 57.80 75.43 P212121 2.0 Takahashi et al (1989)... 

Fig. 26.1 Stereoview showing a small portion of an 1.8 A electron-density map of porcine pancreatic elastase contoured at Ict (1 standard deviation) above mean. The electron-density map, drawn in thin lines, is the direct experimental result while the atomic model, drawn in thick lines, has been built into the electron-density map by the crystallographers. (To learn how to view stereoimages, see Further Reading).

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