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)

Papain [EC 3.4.22.2] was photooxidized using methylene blue as a sensitizer. The photooxidzed enzyme lost its caseinolytic activity and had significantly decreased histidine and tryptophan contents. The tyrosine content was the same before and after the photooxidation. The SH content of the photooxidized enzyme, as determined after reduction with dithiothreitol, was also unchanged. The loss of histidine was always slower than the loss of enzymatic activity, being less than one residue per molecule even when the enzymatic activity was completely lost. However, the inactivation and the oxidation of a histidine residue were pH-dependent in a similar fashion in the pH range of 5.0-8.0, the pH profiles conforming to theoretical titration curves with apparent pKa values of 6.6 and 6.7, respectively. The fact that the ionization of a histidine residue in papain has a normal imidazole pKa value is entirely in accord with the finding for stem bromelain [EC 3.4.22.4] (Murachi, T., Tsudzuki, T., & Okumura, K. (1975) Biochemistry 14, 249-255), and is of great significance in relation to the mechanism of catalysis by these enzymes.
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PMID:Photooxidation of histidine and tryptophan residues of papain in the presence of methylene blue. 23 35

Asparagine-linked oligosaccharides of stem bromelain glycopeptides were quantitatively released by digestion with the almond glycopeptidase which cleaves beta-aspartylglycosylamine linkage in glycopeptides with oligopeptide moieties. The primary structures of the two oligosaccharide components, (Man)3(Xyl)1(Fuc)1(GlcNAc)2 and (Man)2-(Xyl)1(Fuc)1(GlcNAc)2 were elucidated as Man alpha 1 leads to 6Man alpha 1 leads to 6[Xyl beta 1 leads to 2]Man beta 1 leads to 4GlcNAc beta 1 leads 4[Fuc alpha 1 leads to 3]GlcNAc and Man alpha 1 leads to 6[Xyl beta 1 leads to 2]Man beta 1 leads to 4 GlcNAc beta 1 leads to 4[Fuc alpha 1 leads to 3] GlcNAc, respectively.
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PMID:Complete structure of the carbohydrate moiety of stem bromelain. An application of the almond glycopeptidase for structural studies of glycopeptides. 50 Jun 6

A new type of glycopeptidase hydrolyzing beta-aspartylglycosylamine linkages was partially purified from almond emulsin by chromatography on Sephadex G-200 and DE 52. The enzyme degraded stem bromelain glycopeptide, Asn-Asn(Man3,Xyl1,Fuc1,GlcNAc2)-Glu-Ser-Ser, to yield equimolar amounts of intact oligosaccharide, peptide (Asn-Asp-Glu-Ser-Ser), and ammonia. The Km value for the stem bromelain glycopeptide was 4 mM, and the optimum pH was 5.2. The enzyme was markedly inhibited by 10 mM Cu2+, Fe3+, and Zn2+. Thiol inhibitors and actinomycete protease inhibitors had no effect. The glycopeptides used as substrates were prepared from stem bromelain, ovalbumin or ovotransferrin. The enzyme hydrolyzed glycopeptides with 3-11 amino acid residues, whereas it did not hydrolyze glycopeptides with 1-2 amino acid residues. Furthermore, Asn-oligosaccharide was not inhibitory to the enzyme.
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PMID:Some characteristics of a new glycopeptidase acting on aspartylglycosylamine linkages. 73 97

Fruit bromelain FA2, the main proteinase component of the juice of pineapple fruit, has been purified and characterized. 1. Efficient extraction of this enzyme from the crude material was possible using "Cellulosin AP," a microbial polysaccharidase preparation containing cellulase, hemicellulase, and pectinase. The enzyme was purified mainly by successive applications of anion-exchange chromatography, yielding an apparently homogeneous protein as judged by several physical, chemical, and immunochemical criteria. Properties of FA2 include: molecular weight, 31,000; isoelectric point, pH 4.6; absorbance at 280 nm of a 1% solution at pH 7.0 per cm, 19.2. 2. FA2 gave only alanine phenylthiohydantoin upon amino-terminal group analysis by the Edman procedure. Stepwise degradation yielded the amino-terminal sequence Ala-Val-Pro-Gln-Ser-Ile-Asp-Trp-Arg-Asp-Tyr-Gly-Ala. The amino acid composition of FA2 was not markedly different from that of stem bromelain, except for a much smaller lysine content and a smaller alanine content relative to glycine in FA2. FA2 contained neither amino sugars nor neutral carbohydrates as determined by several methods, so FA2 is not a glycoprotein. 3. By labeling the reactive cysteine residue (CYS) with [14C]iodoacetate, the following partial amino acid sequence has been determined. Asn-Glx-Asn-Pro-Cys-Gly-Ala-CYS.
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PMID:Purification and characterization of a proteinase from pineapple fruit, fruit bromelain FA2. 95 52

Pineapple stem bromelain was photooxidized in the presence of Methylene Blue used as a sensitizer. The essential sulfhydryl group of the enzyme protein rapidly became inaccessible to react with 5,5'-dithiobis(nitrobenzoic acid), but the reactivity was readily regained to the original level upon treatment with dithiothreitol. Even after such reduction, the photooxidized enzyme showed a markedly decreased hydrolytic activity on casein. Spectral examination revealed that the oxidized enzyme had tyrosine residues intact. Amino acid analysis showed significant decreases in histidine, ethionine, and tryptophan residues. Photoinactivation occurred in a similar manner also in the presence of tetrathionate which reversibly blocked the essential sulfhydryl group. It is concluded that the irreversible photoinactivation of stem bromelain must be related to the oxidation of histidine, methionine, and tryptophan residues. When the photooxidation was carried out a different pH values ranging from 4.0 to 8.3, the inactivation and the decrease in histidine content were found to be markedly pH dependent. Thus, the photooxidation experiment provided a method for directly measuring the apparent pKa of the ionization of the single histidine residue in stem bromelain. Apparent pKa values of 6.4 and 7.1 were obtained for the histidine imidazole in the absence and in presence of tetrathionate, respectively. In view of these normal pKa values for an imidazole, a mechanism of ionization of the active-site group in a plant thiol proteinase is proposed, in which the validity of mechanism involving a close electronic interaction between histidine and cysteine residues is seriously questioned.
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PMID:Photosensitized inactivation of stem bromelain. Oxidation of histidine, methionine, and tryptophan residues. 112 Jan

Close to 15% of the karatasin proteinase activity in the fruit juice of Bromelia plumieri (karatas) is present outside dialysis Visking tubing in 7 days in 0.2 M acetate buffer (pH) 3.5 or 6.5) containing phenyl mercuric acetate. The small proteinase(s), distinct from the 85% activity in juice due to nondialysable karatasin with a reported Mr of 24,868, separates across Spectrapore (13 kDa) membranes but not across Spectrapore with 3.5 kDa average pore diameter. The dialyzed proteinase is named karatasin-D (K-D). Purified non-Dialysable karatasin can be dissociated to what seems to be K-D by incubation in a buffer solution, containing SDS and 2-mercaptoethanol with phenyl mercuric acetate, in dialysis experiments for 8 days at room temperature using Spectrapore 13 kDa tubing. Thus, native karatasin in B. plumieri fruit juice seem to be the result of association of 2 small molecular mass K-D subunits, linked together by disulfide bonds and electrostatic forces, in equilibrium with small amounts of free K-D molecules. The amino acid composition and partial sequence of karatasin up to the 14th position from the amino terminus have discrete analogies with papain and with stem bromelain.
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PMID:Subunit structure of karatasin, the proteinase isolated from Bromelia plumieri (karatas). 136 18

To investigate the participation of neuropeptides present in the peripheral endings of primary afferent neurons in the inflammatory response, immunoreactive substance P (iSP), calcitonin gene-related peptide (iCGRP) and neurokinin A (iNKA) levels in the s.c. perfusate, and inflammatory response (edema and plasma protein extravasation) evoked in rat paw by noxious stimulation were determined. The effects of these peptides on plasma protein extravasation in the skin of the hind paw of mice were also examined with the pontamine sky blue protein labelling method. The following results were obtained. 1) Immersion of the rat hind paw for 30 min into hot water adjusted to 47 degrees C led to a marked increase in the release of iSP and iCGRP in the subcutaneous perfusate with the formation of thermal edema. 2) Mechanical stimulation (600 g, 10 min) to the hind paw or electrical stimulation of the saphenous and sciatic nerves (10 V, 2 Hz, 1msec duration, 10 min) evoked the increase of iSP release in the perfusate with plasma protein extravasation. 3) iNKA release was not affected by neither heat nor mechanical stimulation. 4) Intraplantar injection of SP, CGRP and NKA induced plasma protein extravasation, the order of potencies being SP greater than CGRP greater than NKA. The action of SP was antagonized by spantide, an SP antagonist. The injection of CGRP with SP produced a synergistic action on plasma protein extravasation. 5) Neonatal pretreatment with capsaicin, which is known to degenerate small-diameter primary afferent neurons, caused the decrease in amount of iSP and iCGRP released during noxious heat stimulation. 6) Pretreatment with Compound 48/80, or stem bromelain and emorphazone, or des-Arg9-[Leu8]-BK, inhibited the iSP release evoked by noxious heat stimulation. 7) Opioids such as morphine (mu-agonist) and ethylketocyclazocine (kappa agonist) inhibited the heat stimulus-evoked iSP release and thermal edema, and the inhibitory effects were antagonized by pretreatment with their antagonists. 8) Morphine or ethylketocyclazocine or [D-Ala2,D-Leu5]-enkephalin (delta-agonist) inhibited the release of iSP evoked by electrical stimulation of the saphenous and sciatic nerves. These results indicate that SP and CGRP present in peripheral endings of small-diameter primary afferent neurons play an important role in the inflammatory response, and that opioids are involved in the regulation of inflammatory response through the inhibition of SP release.
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PMID:[A pharmacological study of the participation of the peripheral endings of primary afferent neurons in the inflammatory response evoked by heat and mechanical noxious stimulation]. 172 88

The pineapple plant (Ananas comosus) was shown to contain at least four distinct cysteine proteinases, which were purified by a procedure involving active-site-directed affinity chromatography. The major proteinase present in extracts of plant stem was stem bromelain, whilst fruit bromelain was the major proteinase in the fruit. Two additional cysteine proteinases were detected only in the stem: these were ananain and a previously undescribed enzyme that we have called comosain. Stem bromelain, fruit bromelain and ananain were shown to be immunologically distinct. Enzymic characterization revealed differences in both substrate-specificities and inhibition profiles. A study of the cysteine proteinase derived from the related bromeliad Bromelia pinguin (pinguinain) indicated that in many respects it was similar to fruit bromelain, although it was found to be immunologically distinct.
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PMID:The cysteine proteinases of the pineapple plant. 232 70

The inhibitory specificity of a protein from potato tubers that inhibits cysteine proteinases (potato cysteine proteinase inhibitor, PCPI) has been compared with that of chicken egg-white cystatin. Most proteinases that are inhibited by cystatin were also inhibited by PCPI, but the potato inhibitor inhibited stem bromelain and fruit bromelain, which are not inhibited by cystatin, and for which no protein inhibitor of comparable potency has previously been described. In contrast, papaya proteinase IV was unaffected by PCPI as it is by the cystatins, and the exopeptidase, dipeptidyl peptidase I, is inhibited by cystatins, but was unaffected by PCPI. The differences in inhibitory specificity between these proteins may well reflect differences between superfamilies of cysteine proteinase inhibitors.
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PMID:Inhibition of cysteine proteinases by a protein inhibitor from potato. 240 57

To investigate a physiological function of substance P (SP) present in the peripheral ending of sensory neurons, we determined immunoreactive SP (iSP) levels in the s.c. perfusate and the amount of edema evoked in rat paw by noxious heat stimulation. We found that immersion of rat paw into hot water (47 degrees C) for 30 min led to a significant increase of iSP in the perfusate and about 50% increase in paw volume. Neonatal pretreatment with capsaicin inhibited significantly the increase in both iSP and paw volume evoked by noxious heat stimulation. Acute and chronic denervation of the sciatic and saphenous nerves also inhibited the heat-evoked iSP release and edema remarkably. Intraplantar injection of SP evoked an increase in paw volume in dose-dependent manner. This increasing effect of SP on paw volume was more substantial than that produced by histamine. Simultaneous treatment with stem bromelain and emorfazone decreased significantly the heat-evoked iSP release and edema. These results suggest that 1) SP produced by noxious heat stimulation in the periphery may be released from the afferent fibers with small-diameter, 2) bradykinin may intervene in this SP release and 3) SP released in the periphery may be closely related to the edema formation of the thermal injury reaction.
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PMID:Contribution of substance P to heat-induced edema in rat paw. 244 42


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