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

Soybean protein, casein, bonito protein and chicken protein, each as foodstuff protein, were hydrolyzed with four proteinases; namely, pepsin, trypsin, alpha-chymotrypsin and bromelain. Since the chicken protein hydrolysate with bromelain possessed the most favorable umami taste, eleven peptides were isolated from the chicken protein hydrolysate by successive chromatography on ODS, Amberlite IR-120B, Amberlite IRA-410 and AG-50W; their structures were Asp-Ala, Asp-Val, Glu-Glu, Glu-Val, Ala-Asp-Glu, Ala-Glu-Asp, Asp-Glu-Glu, Asp-Glu-Ser, Glu-Glu-Asn, Ser-Pro-Glu, and Glu-Pro-Ala-Asp. Many of them did not show any umami taste by themselves, but Glu-Glu, Glu-Val, Ala-Asp-Glu, Ala-Glu-Asp, Asp-Glu-Glu, and Ser-Pro-Glu were recognized to enhance the umami taste of 0.02% 5'-inosine monophosphate (IMP). A combination of these peptides, especially 0.5% each of Glu-Glu, Glu-Val, Asp-Glu-Glu and Glu-Glu-Asn, with 0.02% IMP produced a delicious "full" umami taste.
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PMID:Isolation of peptides from an enzymatic hydrolysate of food proteins and characterization of their taste properties. 1022 42

A new papain-like cysteine peptidase isolated from fruits of Pseudananas macrodontes (Morr.) Harms, a species closely related to pineapple (Ananas comosus L.), has been purified and characterized. The enzyme, named macrodontain I, is the main proteolytic component present in fruit extracts and was purified by acetone fractionation followed by anion-exchange chromatography. Separation was improved by selecting both an adequate pH value and a narrow saline gradient. Optimum pH range (more than 90% of maximum activity with casein) was achieved at pH 6.1-8.5. Homogeneity of the enzyme was confirmed by bidimensional electrophoresis and mass spectroscopy (MS). Molecular mass of the enzyme was 23,459 (MS) and its isoelectric point was 6.1. The alanine, glutamine, and tyrosine derivatives were strongly preferred when the enzyme was assayed on N-alpha-CBZ-l-amino acid p-nitrophenyl esters. The N-terminal sequence of macrodontain (by comparison with the N-terminus of 30 plant proteases with more than 50% homology) showed a great deal of sequence similarity to the other pineapple-stem-derived cysteine endopeptidases, being 85.7, 85. 2, and 77.8% identical to comosain, stem bromelain, and ananain, respectively. It seems clear that the Bromeliaceae endopeptidases are more closely related to each other than to other members of the papain family, suggesting relatively recent divergence.
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PMID:Purification and characterization of macrodontain I, a cysteine peptidase from unripe fruits of Pseudananas macrodontes (Morr.) harms (Bromeliaceae). 1068 43

1. The purpose of this study was to evaluate the effects of protein source and enzyme supplementation on protein digestibility and chyme characteristics in broilers. 2. One hundred and twenty growing (13 d old) and 60 finishing (34 d old) Arbor Acre strain commercial male broilers were selected and placed into individual metabolic cages. 3. The experiment was a 5 x 2 factorial arrangement with 5 different sources of protein: casein, fish meal, soybean meal (SBM), soy protein concentrate (SPC), maize gluten meal (MGM) and two levels of protease (bromelain), 0 and 65 CDU/kg diets. 4. The diets were iso-nitrogenous and semi-purified, with Cr2O3 as an indicator for determination of ileal digestibility and chyme characteristics. 5. Apparent ileal protein digestibility (AIPD) in both growing and finishing chickens was highest on the casein diet, followed by fish meal, SBM, SPC and MGM. 6. Enzyme inclusion did not improve protein digestibility, but significantly decreased the digesta pH value in the gizzard and increased pH in the ileum in the 3-week-old broilers. 7. The digesta pH values in the gizzard and duodenum were significantly lower in the SBM and fish meal groups compared with the other protein groups. The molecular weight distribution pattern of the soluble protein in the chyme of the gastrointestinal (GI) segments showed a similar trend, regardless of the enzyme inclusion or the stage of growth. 8. The molecular weight profile of soluble protein changed dynamically in the casein fed broilers from the gizzard to ileum and the low molecular weight proteins, < 7 kDa, reached maximum levels at the ileum. The molecular weight profile of the soluble protein in the SBM and SPC changed between the jejunum and the ileum and in the intermediate molecular soluble protein weight (7 to 10 kDa) was significantly decreased. This indicated that the hydrolysis process began from the middle to the posterior end of the small intestine. 9. Similar profiles were also shown with fish meal protein. The pattern of distribution, however, did not show any prominent change in the GI segments of the MGM group. 10. The pepsin, trypsin and chymotrypsin protease activity in the gizzard and duodenum were highest in the casein group and lowest in the MGM group as compared with the other protein groups. 11. The rate change in the patterns of molecular weight distribution in soluble protein and the digestive enzyme activity provide indications of the partial digestibility of different protein sources. The exogenous enzyme, bromelain, did not show any beneficial effect on protein digestion.
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PMID:Effects of sources of protein and enzyme supplementation on protein digestibility and chyme characteristics in broilers. 1219 2

A serine protease, subtilisin YaB, produced by alkalophilic Bacillus YaB, shows promises as a potent meat tenderizer, because its substrate specificity is for small amino acids, which are found at high levels in meat connective tissue proteins. Substrate specificity engineering of the substrate binding pockets was used to generate more suitable meat-tenderizing mutants, G124A, G124V, G159A, and G159S, derived from recombinant wild subtilisin YaB and expressed in Bacillus subtilis DB104. The characteristics of these recombinant enzymes were studied to evaluate their usefulness as improved meat tenderizers. The proteolytic activities of recombinant subtilisin YaB, engineered subtilisin YaBs, and commercially available papain, bromelain, collagenase, and elastase were compared using elastin, collagen, casein, and myofibrillar proteins as substrates. Hydrolysis of beef proteins was evaluated using the myofibrillar fragmentation index and collagen solubility. The results demonstrated that recombinant mutant G159A was the most improved meat tenderizer and can be used in the meat pH range of 5.5-6.0 and the temperature range of 10-50 degrees C. Contrary to the result obtained from artificial substrate, mutant enzymes engineered on G124 residues did not exhibit better tenderizing ability when elastin, collagen, or meat was used as substrate, suggesting the necessity of evaluation by real substrate before protein-engineered enzymes are applied commercially.
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PMID:Application potency of engineered G159 mutants on P1 substrate pocket of subtilisin YaB as improved meat tenderizers. 1235 2

The present work reports the characterization of Fastuosain, a novel cysteine protease of 25kDa, purified from the unripe fruits of Bromelia fastuosa, a wild South American Bromeliaceae. Proteolytic activity, measured using casein and synthetic substrates, was dependent on the presence of thiol reagents, having maximum activity at pH 7.0. The present work reports cDNA cloning of Fastuosain; cDNA was amplified by PCR using specific primers. The product was 1096pb long. Mature fastuosain has 217 residues, and with the proregion has a total length of 324 residues. Its primary sequence showed high homology with ananain(74%), stem bromelain (66%) and papain (44%).
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PMID:Preliminary functional characterization, cloning and primary sequence of Fastuosain, a cysteine peptidase isolated from fruits of Bromelia fastuosa. 1645 75

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

Bromelain is a basic, 23.8 kDa thiol proteinase obtained from stem of the pineapple plant (Ananas comosus) and is unique in containing a single oligosaccharide chain attached to the polypeptide. This property allowed its affinity binding and favorable orientation on a Sepharose support pre-coupled with the lectin, concanavalin A (Con A). For comparison, bromelain was also immobilized by covalently coupling to the CNBr-activated Sepharose. The preparation obtained was more resistant to thermal inactivation as evident from the retention of over 50% activity after incubation at 60 degrees C for 100 min (as compared to 20% retained by the native enzyme and 30% retained by the covalently immobilized enzyme), exhibited a broader pH-activity profile with the enzyme retaining over 60% activity at pH 11 (as compared to over 25% retained by native and the enzyme immobilized covalently). The native, covalently-coupled and affinity-bound bromelains had apparent K (m) values of 1.1, 2 and 0.54 mg/ml, respectively using casein as the substrate. The V (max) values remained unaffected on immobilization.
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PMID:Bioaffinity based oriented immobilization of stem bromelain. 1678 78

Stem bromelain was covalently coupled to a thermosensitive polymer of N-isopropylacrylamide (p(NIPAm)) either through the amino groups of the enzyme (randomly coupled) or via the lone oligosaccharide chain (uniformly coupled). The enzyme coupled via the oligosaccharide chain exhibited better access to the substrate casein as compared to the preparation in which the amino groups formed the point of contact between the enzyme and the polymer. Native bromelain exhibited a pH optimum of 8.0 and a broad pH-activity profile. The polymer-coupled preparations exhibited broader pH-activity profiles and shifting of pH optimum to 10.0 at 35 degrees C. At 25 degrees C, the shifting of pH optimum was observed for the randomly coupled enzyme only. The temperature-activity profiles of bromelain coupled to p(NIPAm) also showed appreciable broadening and the preparations retained greater fraction of maximum activity above the temperature optimum. The optimum temperature of the uniformly oriented preparation also rose to 70 degrees C. Inactivation rates of the polymer-coupled bromelain were remarkably low at 60 degrees C as compared to the native protease, and binding of antibromelain antibodies improved the resistance to inactivation of the polymer-coupled preparations. The cleavage patterns of hemoglobin and IgG by the native bromelain and the polymer-coupled preparations were comparable.
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PMID:Additional stabilization of stem bromelain coupled to a thermosensitive polymer by uniform orientation and using polyclonal antibodies. 1744 84

A heavy-metal assay has been developed using bromelain, a protease. The enzyme is assayed using casein as a substrate with Coomassie dye to track completion of hydrolysis of casein. In the absence of inhibitors, casein is hydrolysed to completion, and the solution is brown. In the presence of metal ions such as Hg2+ and Cu2+, the hydrolysis of casein is inhibited, and the solution remains blue. Exclusion of sulfhydryl protective agent and ethylenediaminetetraacetic in the original assay improved sensitivity to heavy metals several fold. The assay is sensitive to Hg2+ and Cu2+, exhibiting a dose-response curve with an IC50 of 0.15 mg 1(-1) for Hg2+ and a one-phase binding curve with an IC50 of 0.23 mg 1(-1) for Cu2+. The IC50 value for Hg2+ is found to be lower to several other assays such as immobilized urease and papain assay, whilst the IC50 value for Cu2+ is lower than immobilized urease, 15-min Microtox, and rainbow trout.
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PMID:An inhibitive determination method for heavy metals using bromelain, a cysteine protease. 1855 17

Lupin seed gamma-conglutin, orally administered to animal models, has been shown to display glucose-controlling properties. Therefore, we have addressed the study of gamma-conglutin susceptibility to proteolytic enzymes in vitro as the basis to unveil its metabolic fate in the body. Pepsin treatment at pH 2.0 and 3.0 caused extensive proteolytic breakdown, while at pH 4.0, where pepsin is minimally active, gamma-conglutin was unaffected. Aliquots of the pepsin-treated protein were further incubated with pancreatin at neutral pH. If the protein backbone was already cleaved by pepsin action, then the breakdown by pancreatin was almost complete; alternatively, pancreatin did not affect at all gamma-conglutin polypeptide chain. This was not due to an inhibitory activity of gamma-conglutin, because co-incubation with casein showed complete breakdown of the milk protein. Furthermore, gamma-conglutin was incubated with bromelain, a proteinase effective between pH 4.0 and 7.0. A sharp transition from the uncleavable to the fully cleavable form of gamma-conglutin was observed below pH 4.25. Therefore, it was concluded that (i) gamma-conglutin is resistant to proteolysis at pH greater than 4.0, likely because of a compact native conformation, (ii) an acidic pH renders the protein susceptible to proteases, suggesting the occurrence of a trans conformation, which has also been observed by circular dichroism spectral analysis, and (iii) the protein undergoes an "all or none" degradation pathway, regardless of the enzyme used.
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PMID:Susceptibility of lupin gamma-conglutin, the plasma glucose-lowering protein of lupin seeds, to proteolytic enzymes. 1970 31


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