Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.22.32 (bromelain)
 1,025 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The sensitivity of highly purified human fibroblast interferon and partially purified human leukocyte interferon to several proteolytic and glycolytic enzymes was determined with respect to antiviral activity, isoelectric point, molecular weight, and thermal stability. Leucine aminopeptidase altered the distribution of isoelectric points for both interferons but produced little change in molecular weights; this enzyme somewhat reduced the activity of only leukocyte interferon. Treatment of fibroblast interferon with carboxypeptidases A and B did not greatly decrease antiviral activity, but it did slightly reduce the molecular weight of the interferon and substantially altered the distribution of isoelectric point values; similar treatment of leukocyte interferon caused some loss in activity, especially of the 17,000-molecular-weight species. Both interferons were inactivated rapidly by treatment with the endoproteases trypsin, pepsin, bromelain, and subtilisin. Chymotrypsin shifted the isoelectric points of both interferons, but only leukocyte interferon was significantly inactivated. Treatment with neuraminidase and beta-galactosidase changed the isoelectric point distribution but did not affect the activity or thermal stability of either interferon; such a treatment reduced the molecular weight of fibroblast interferon and the size heterogeneity of leukocyte interferon. Treatment with neuraminidase and then leucine aminopeptidase greatly reduced the activity of both interferons, especially leukocyte interferon. The data indicate that biologically active forms of fibroblast and leukocyte interferons can be distinguished by their relative sensitivity to certain proteases.
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PMID:Enzymatic modifications of human fibroblast and leukocyte interferons. 616 Feb 60

A new method for removing nearly all active endoproteinases from fluids called "sandwich affinity chromatography" is described. It is based on strong chelate binding of alpha 2-macroglobulin (alpha 2M) and its proteinase complexes to Zn2+-bis-carboxymethylamino-Sepharose (Zn chelate-Sepharose) and its ability to complex most active endoproteinases. The preferred performance minimizing unspecific protein adsorption is binding first alpha 2M to Zn chelate-Sepharose and then adsorbing the proteinase to the alpha 2M-Zn chelate-Sepharose using elevated salt concentrations. A suitable standard buffer, in which most proteases and alpha 2M are active and the protease-alpha 2M complex remains bound to Zn chelate-Sepharose, is 0.02 mol/liter sodium phosphate, pH 6.5, containing 0.15 mol/liter NaCl. As an example, the reaction of trypsin with alpha 2M-Zn chelate-Sepharose was studied. After saturating Zn chelate-Sepharose first with alpha 2M and then with trypsin under standard conditions, the bound alpha 2M equals the bound trypsin activity (measured with Chromozym TRY). The specific binding capacity of alpha 2M-Zn chelate-Sepharose for proteases was determined in this way to be 30-40 U trypsin, i.e., 0.40-0.54 mg/ml of gel. The balance and the fact that the bound trypsin is inaccessible to soybean trypsin inhibitor indicate that at these conditions no unspecific trypsin binding occurs. Chymotrypsin, thermolysin, elastase, bromelain, ficin, and papain are also bound at standard conditions but not exoproteases like carboxypeptidases A and Y. Advantages of the sandwich affinity chromatography are the simple loading procedure by adsorption, the high capacity of the gel material, and the possibility to reuse the Zn chelate-Sepharose after eluting reacted alpha 2M and reloading with new alpha 2M.
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PMID:Removal of endoproteinases from biological fluids by "sandwich affinity chromatography" with alpha 2-macroglobulin bound to zinc chelate-Sepharose. 620 48

A novel total enzymatic synthesis of [Leu]- and [Met]-enkephalin derivatives was accomplished in low-water content systems at a preparative scale. alpha-Chymotrypsin, papain, thermolysin and bromelain adsorbed on Celite were used as catalysts. Organic solvents such as acetonitrile and ethyl acetate with small amounts of buffer added or at specific water activity were used as reaction media. Simple readily available amino acid ester derivatives were used as starting building blocks. This feature allowed the possibility of using the products in one step directly as acyl-donor ester, without any chemical or enzymatic modification, in the next enzymatic coupling. The optimal strategy for the synthesis of the enkephalin derivatives was different depending on the carboxy terminal group. The preparation of the carboxy-terminal amide derivatives (R-Tyr-Gly-Gly-Phe-Leu[Met]-NH2) was achieved via 4 + 1 fragment condensation catalyzed by alpha-chymotrypsin. The carboxy-terminal ethyl ester derivatives (R-Tyr-Gly-Gly-Phe-Leu[Met]-OEt) were obtained via 2 + 3 condensation catalyzed by bromelain, a quite unusual protease for peptide synthesis but more effective than papain in this coupling. Both syntheses were carried out in four enzymatic steps and one or two chemical deprotection steps routinely used in peptide synthesis. The overall yields of pentapeptide derivatives were between 40-54% of pure product.
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PMID:Enzymatic peptide synthesis in low water content systems: preparative enzymatic synthesis of [Leu]- and [Met]-enkephalin derivatives. 760 86

The proteolytic activities of alpha-chymotrypsin, trypsin, pepsin, bromelain, and an extract from germinating pumpkin seeds (Cucurbita moschata) were determined by their ability to effect the release of 1-anilino-8-naphthalenesulfonate bound to internal hydrophobic sites in intact protein substrates. Casein, glyceraldehyde-3-P dehydrogenase, urease, catalase, pumpkin seed globulin, and bovine serum albumin enhanced the fluorescence of 1-anilino-8-naphthalenesulfonate sufficiently to be used as proteolytic substrates. Chymotrypsin, trypsin, pepsin, and bromelain exhibited activity against all or almost all of the protein substrates. The activity of 1 mug of alpha-chymotrypsin or trypsin and 100 ng of pepsin could be easily detected by this method of assay within 4 to 5 minutes depending upon the substrate. The enzyme extracted from 3-day germinated pumpkin seeds exhibited strong activity only against pumpkin seed globulin, weak activity against the globulins of squash and cucumber and casein, and no activity against the other protein substrates. Activity against pumpkin globulin was maximal at pH 7.4. When assayed by an increase in ninhydrin-positive products, the enzyme extract from pumpkin seeds also showed strong activity against pumpkin globulin and weak activity against casein. The 1-anilino-8-naphthalenesulfonate-fluorescence method was at least 20 times more sensitive than the ninhydrin method and was 10 to 20 times more rapid.
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PMID:Globulin-specific Proteolytic Activity in Germinating Pumpkin Seeds as Detected by a Fluorescence Assay Method. 1665 2