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 saline extract of Bauhinia bauhinioides dry seeds was shown to inhibit cruzipain, a cysteine proteinase from Trypanosoma cruzi. The inhibitory activity was assigned to a protein with 164 amino acid residues and molecular mass of 18 034 Da that was purified by chromatography on DEAE-Sephadex, trypsin-Sepharose (removal of trypsin inhibitors), Mono Q and a reversed-phase C4 column. The primary structure is homologous to other plant Kunitz-type inhibitors, but it lacks cysteine residues and therefore the disulfide bridges. No methionine residue was identified by amino acid sequencing. The inhibition of cruzipain fits into a slow-tight binding mechanism with a low dissociation constant (Ki 1.2 nM). The studied Bauhinia protein also inhibits cruzain (Ki 0.3 nM), a C-terminally truncated recombinant species of cruzipain. Cathepsin L, a cysteine proteinase with high homology to cruzipain, is also inhibited (Ki 0.22 nM), but not cathepsin B, papain, bromelain or ficin.
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PMID:Structure of cruzipain/cruzain inhibitors isolated from Bauhinia bauhinioides seeds. 1151 40

Kiwifruit cysteine proteinase inhibitors (KCPIs) were purified from the cortex and seeds of kiwifruit after inactivation of the abundant cortex cysteine proteinase actinidain. One major (KCPI1) and four minor cystatins were identified from Actinidia deliciosa ripe mature kiwifruit cortex as well as a seed KCPI from A. chinensis. The predominant cortex cystatin, KCPI1, inhibited clan CA, family C1 (papain family) cysteine proteinases (papain, chymopapain, bromelain, ficin, human cathepsins B, H and L, actinidain and the house dust mite endopeptidase 1), while cysteine proteinases belonging to other families, [clostripain (C11), streptopain (C10) and calpain (C2)] were not inhibited. Inhibition constants (K(I)) ranged between 0.001 nM for cathepsin L and 0.98 nM for endopeptidase 1. The K(I) (14 nM) for KCPI1 inhibiting actinidain is at least 2 orders of magnitude higher than for other plant proteinases measured. The cortex KCPI1 and a seed KCPI purified from seeds had the same N-terminal sequence (VAAGGWRPIESLNSAEVQDV). BLAST-matching the peptide sequence against an in-house generated Actinidia EST database, identified 81 cDNAs that exactly matched the measured KCPI1 peptide sequence. Peptide sequences of two other cortex KCPIs each exactly matched a predicted peptide sequence of a cDNA from kiwifruit. The predicted peptide sequence of KCPI1 of 116 amino acids encodes a signal peptide and does not contain cysteine. Without the signal peptide (mature protein), KCPI1 has a molecular mass of approximately 11 kDa, possesses the consensus sequence characteristic for the phytocystatins and shows the highest homology to a cystatin from Citrusxparadisi (52% identity). This is the first report of phytocystatins from the Ericales.
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PMID:Purification and characterization of phytocystatins from kiwifruit cortex and seeds. 1469 68

Many plants contain latex that exudes when leaves are damaged, and a number of proteins and enzymes have been found in it. The roles of those latex proteins and enzymes are as yet poorly understood. We found that papain, a cysteine protease in latex of the Papaya tree (Carica papaya, Caricaceae), is a crucial factor in the defense of the papaya tree against lepidopteran larvae such as oligophagous Samia ricini (Saturniidae) and two notorious polyphagous pests, Mamestra brassicae (Noctuidae) and Spodoptera litura (Noctuidae). Leaves of a number of laticiferous plants, including papaya and a wild fig, Ficus virgata (Moraceae), showed strong toxicity and growth inhibition against lepidopteran larvae, though no apparent toxic factors from these species have been reported. When the latex was washed off, the leaves of these lactiferous plants lost toxicity. Latexes of both papaya and the wild fig were rich in cysteine-protease activity. E-64, a cysteine protease-specific inhibitor, completely deprived the leaves of toxicity when painted on the surface of papaya and fig leaves. Cysteine proteases, such as papain, ficin, and bromelain, all showed toxicity. The results suggest that plant latex and the proteins in it, cysteine proteases in particular, provide plants with a general defense mechanism against herbivorous insects.
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PMID:Papain protects papaya trees from herbivorous insects: role of cysteine proteases in latex. 1473 Dec 57

Promastigotes of all pathogenic Leishmania species secrete acid phosphatase (SAcP) activity during their growth in vitro. It has been suggested that this enzyme may play a role in the survival of the parasite within its sandfly-vector host. To carry out such functions, SAcP would have to be relatively resistant to endogenous sandfly gut-proteases. Therefore, the current study was undertaken to ascertain whether L. donovani SAcP activity was affected by treatment with various proteases. Native L. donovani SAcP was treated with a variety of serine-, thiol-, metallo- and mixed-proteases and subsequently assayed for enzymatic activity. Of the eleven proteases tested, only bromelain and subtilisin treatments caused a pronounced reduction in SAcP activity. Treatment of SAcP with seven out of the remaining nine proteases, resulted in an overall enhancement in SAcP enzymatic activity ranging from approximately 10% (e.g. with trypsin) to > or = 90% (e.g. with ficin). The resistance of the Leishmania SAcP to various proteases may prolong its functional life within the sandfly gut and help to facilitate parasite infection in this host.
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PMID:The human pathogen Leishmania donovani secretes a histidine acid phosphatase activity that is resistant to proteolytic degradation. 1506 72

The cDNA of a cystein peptidase inhibitor was isolated from sugarcane and expressed in Escherichia coli. The protein, named canecystatin, has previously been shown to exert antifungal activity on the filamentous fungus Trichoderma reesei. Herein, the inhibitory specificity of canecystatin was further characterized. It inhibits the cysteine peptidases from plant source papain (Ki =3.3nM) and baupain (Ki=2.1x10(-8)M), but no inhibitory effect was observed on ficin or bromelain. Canecystatin also inhibits lysosomal cysteine peptidases such as human cathepsin B (Ki=125nM), cathepsin K (Ki=0.76nM), cathepsin L (Ki=0.6nM), and cathepsin V (Ki=1.0nM), but not the aspartyl peptidase cathepsin D. The activity of serine peptidases such as trypsin, chymotrypsin, pancreatic, and neutrophil elastases, and human plasma kallikrein is not affected by the inhibitor, nor is the activity of the metallopeptidases angiotensin converting enzyme and neutral endopeptidase. This is the first report of inhibitory activity of a sugarcane cystatin on cysteine peptidases.
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PMID:Inhibitory selectivity of canecystatin: a recombinant cysteine peptidase inhibitor from sugarcane. 1524

Proteases regulate numerous biological processes with a degree of specificity often dictated by the amino acid sequence of the substrate cleavage site. To map protease/substrate interactions, a 722-member library of fluorogenic protease substrates of the general format Ac-Ala-X-X-(Arg/Lys)-coumarin was synthesized (X=all natural amino acids except cysteine) and microarrayed with fluorescent calibration standards in glycerol nanodroplets on glass slides. Specificities of 13 serine proteases (activated protein C, plasma kallikrein, factor VIIa, factor IXabeta, factor XIa and factor alpha XIIa, activated complement C1s, C1r, and D, tryptase, trypsin, subtilisin Carlsberg, and cathepsin G) and 11 papain-like cysteine proteases (cathepsin B, H, K, L, S, and V, rhodesain, papain, chymopapain, ficin, and stem bromelain) were obtained from 103,968 separate microarray fluorogenic reactions (722 substrates x 24 different proteases x 6 replicates). This is the first comprehensive study to report the substrate specificity of rhodesain, a papain-like cysteine protease expressed by Trypanasoma brucei rhodesiense, a parasitic protozoa responsible for causing sleeping sickness. Rhodesain displayed a strong P2 preference for Leu, Val, Phe, and Tyr in both the P1=Lys and Arg libraries. Solution-phase microarrays facilitate protease/substrate specificity profiling in a rapid manner with minimal peptide library or enzyme usage.
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PMID:High throughput substrate specificity profiling of serine and cysteine proteases using solution-phase fluorogenic peptide microarrays. 1570 70

The development of a simultaneous multiple substrate enzymatic assay based on electrospray ionization mass spectrometry (ESI-MS) detection is described. This multiplexing assay scheme was employed in a parallel proteolytic enzyme activity screening. As model systems, the respective activities of trypsin, thrombin, chymotrypsin, bromelain, ficin and elastase towards seven different substrates were assessed. The resulting activity patterns were evaluated semi-quantitatively ranking the enzymatic activities in five classes of activity (very high, high, medium, low and no activity) with respect to the individual substrates. The validity of the MS-based multiplexing assay scheme was proved by comparison with the results obtained from single substrate assays detected by means of UV/vis absorption at 405 nm, showing good agreement of the resulting activity patterns and classifications.
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PMID:Assessing protease activity pattern by means of multiple substrate ESI-MS assays. 1591 32

High molecular weight kininogen (HMWK) and low molecular weight kininogen (LMWK) have been purified from sheep (Avis Arias) plasma in three steps involving ammonium sulphate precipitation, column chromatography on Sephacryl-300HR and ion exchange chromatography on DEAE cellulose. HMWK gave a single band on native and SDS-PAGE with a molecular weight corresponding to 280 kDa. Under reducing conditions purified HMWK was again resolved to a single band with molecular weight corresponding to 140 kDa indicative of its dimeric nature. LMWK was resolved into two isoforms named as LMWK1 and LMWK2, with an apparent molecular weight of 68 kDa. The yield of HMWK, LMWK1 and 2 was about 8.1, 5.63 and 10.65 respectively. HMWK, LMWK1 and 2 strongly inhibited activities of ficin and papain but not of trypsin, chymotrypsin and bromelain. Ki values estimated for HMWK with papain and ficin was 0.8 and 0.6 nM respectively. Ki values estimated for LMWK1 and 2 with papain were 2.40 and 2.00 nM respectively. Binding of HMWK, LMWK1 and 2 to activated papain were accompanied by pronounced changes in secondary and tertiary structure that are compatible with perturbations of environment of aromatic residues.
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PMID:Purification and characterization of kininogens from sheep plasma. 1600 51

A unique 33-kDa cysteine protease (Mir1-CP) rapidly accumulates at the feeding site in the whorls of maize (Zea mays L.) lines that are resistant to herbivory by Spodoptera frugiperda and other lepidopteran species. When larvae were reared on resistant plants, larval growth was reduced due to impaired nutrient utilization. Scanning electron microscopy (SEM) indicated that the peritrophic matrix (PM) was damaged when larvae fed on resistant plants or transgenic maize callus expressing Mir1-CP. To directly determine the effects of Mir1-CP on the PM in vitro, dissected PMs were treated with purified, recombinant Mir1-CP and the movement of Blue Dextran 2000 across the PM was measured. Mir1-CP completely permeabilized the PM and the time required to reach full permeability was inversely proportional to the concentration of Mir1-CP. Inclusion of E64, a specific cysteine protease inhibitor prevented the damage. The lumen side of the PM was more vulnerable to Mir1-CP attack than the epithelial side. Mir1-CP damaged the PM at pH values as high as 8.5 and more actively permeabilized the PM than equivalent concentrations of the cysteine proteases papain, bromelain and ficin. The effect of Mir1-CP on the PMs of Helicoverpa zea, Danaus plexippus, Ostrinia nubilalis, Periplaneta americana and Tenebrio molitor also was tested, but the greatest effect was on the S. frugiperda PM. These results demonstrate that the insect-inducible Mir1-CP directly damages the PM in vitro and is critical to insect defense in maize.
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PMID:Degradation of the S. frugiperda peritrophic matrix by an inducible maize cysteine protease. 1624 50

Two cysteine proteinase inhibitors I and II were purified from goat kidney using alkaline denaturation, ammonium sulphate fractionation, gel filtration on Sephadex G-75 and ion exchange chromatography on DEAE cellulose. The purified inhibitors were homogenous and showed a single band on SDS PAGE under reducing and non-reducing conditions with an apparent molecular mass of 67 kDa. The cystatin forms were stable in the range of pH 3-10 and up to 95 degrees C. Immunological identity with the sheep LMW kininogen was obtained suggesting that the inhibitor is closely related to kininogens. Spectral studies confirm that the inhibitors have predominantly an alpha-helical structure and undergo major conformational changes during complex formation with papain. The inhibitors had similar inhibitory activities on cysteine proteinases. Both inhibitors inhibited papain, ficin and bromelain competitively, with maximum affinity for papain. The overall lower affinity of these inhibitors to cysteine proteinases compared to other known cystatins can be attributed to the unusual N-terminal sequence where Leu is substituted by Ile. Furthermore, N-terminal sequence analysis revealed maximum homology to mammalian LMW kininogen.
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PMID:Isolation, characterization and kinetics of goat cystatins. 1625 55


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