Cysteine proteases bromelain

Cysteine proteases are a class of enzymes that have been widely studied over the years. The overall principles of substrate recognition, catalysis, and inhibition are now reasonably well documented. This enzyme class includes the plant proteases such as papain, actinidin, and bromelain, and several mammalian lysosomal cathe-psins. By far the majority of the literature reports dealing with cysteine proteases describe results obtained with the enzyme papain, because it is considered to be the archetype of this enzyme class. 

Cysteine proteases are so called because of a critical cysteine involved (together with an adjacent histidine) in the catalytic mechanism. Cysteine proteases include papain-related proteases, calpain-related proteases and the caspases. Papain-like cysteine proteases include the plant enzymes actinidin, aleurain, bromelain, caricain, chymopapain, ficin and papain and the lysosomal cathepsins B, C, H, K, L and S. Cathepsin C is multimeric (MW -200,000), but the other papain-related proteases are monomeric with MWs of about 20,000-35,000. While cathepsin C is a dipeptidyl aminopeptidase, the other enzymes are endopeptidases. Cathepsin B is an endopeptidase and a dipeptidyl carboxypeptidase. Cathepsin H is an endopeptidase and an aminopeptidase. In higher animals, cathepsin B generates peptides from antigens for presentation to T cells by the major histocompatibility... 

Bromelain is a mixture of cysteine proteases obtained from pineapple stems (Ananas comosus, Bromeliaceae) that has been used therapeutically for the treatment of inflammation and trauma [119]. 7n vitro, it has varied stimulatory effects on leukocyte populations, increases CD2-mediated T cell activation, enhances Ag-independent binding to monocytes, etc. The effects of bromelain have previously been attributed to its degradative action at cell surfaces. However, it also acts independent of the removal of cell surface molecules [120]. In order to investigate the possible hormonelike effects of bromelain on intracellular signalling, its effects on TCR7CD3 signalling and IL-2 production were studied. It was observed that bromelain inhibits ERK-2 activation in ThO cells stimulated via the TCR, or with combined TPA plus calcium ionophore. In addtion, bromelain decreased IL-2, IFN-y, and IL-4 mRNA accumulation in ThO cells stimulated with TPA plus calcium ionophore, while the cytokine mRNA accumulation in cells stimulated via TCR was not affected. It seems that bromelain does not act on ERK-2 directly but also inhibits p2r activation, an effector molecule upstream from ERK-2 in the Raf-1/MEKl/ERK kinase cascade. Since p21 is an effector for multiple MAPK pathways, it is likely that bromelain affects other MAPK signalling cascades, such as the INK pathway or p38 MAPK pathway [121],... 

For soluble and immobilized bromelain, temperature increase is accompanied by a decrease in residual enzyme activity. A more complex form of denaturation occurs with the immobilized enzyme, which may involve a two-phase process. Immobilization offers more resistance to denaturation at the higher temperature of 60°C where the second phase is prolonged by a factor of three [60]. Differential scanning calorimetry experiments showed that bromelain is an exceptional protease among the cysteine proteases, illustrated by the fact that its thermal denaturation is consistent with an irreversible two-state model [61]. Also, the far UV circular dichroism spectrum of bromelain differs from those of papain and chymopapain and therefore represents a third spectral class within the cysteine proteinase family [62],... 

Albumin binds OPs on tyrosine 411 of human albumin or tyrosine 410 of bovine albumin (Sakurai ei aL, 2004), The adjacent arginine residue activates this tyrosine for reaction with esters as well as with OPs. Other non-serinc hydrolases that react with OPs include papain and bromelain, which are cysteine proteases that stochiometrically bind OPs on tyrosine, without loss of protease activity (Murachi. 1963 Chaiken and Smith, 1969). 

Fig. 4. (a) dual function of catalytic triad in serine proteases, where catalysis is performed (i) by the charge relay system, and (ii) by general acid-general base (the oxyanion hole is formed by and Si residues), (b) fimction and role of the catalytic residues C s, H , D158 and/ or Ni s (j bromelain) in cysteine proteases of papain family a hydrogen bond... 

Corresponding equations were derived for the cysteine proteases ficin, actinidin, bromelain B and D, and the serine proteases subtilisin, chymotrypsin, and trypsin. In all cases, the coefficient of the electronic a term is around 0.4-0.7. Whereas the other regression coefficients in equations (16) and (17) are also relatively similar, the constant terms of both equations differ by about 1.8 log units, which indicates that the lower lipophilicity of the mesyl group, as compared with a much more lipophilic benzoyl group, is responsible for the lower binding affinities of the mesyl amides. To prove this hypothesis, a series of (4-X-Phe)-CONHCH2COO-pyrid-3-yl analogs 12 was synthesized and tested. As expected, lipophilicity determines the structure-activity relationship in this part of the molecules (equation IS). " " ... 

Enzymes from plants have been used since andent times, even though their nature remained obscure until the nineteenth century. Malting of cereals and the hydrolysis of complex polysaccharides are two of the oldest uses of enzymes in human history. Like their animal counterparts, most commerdally available plant-derived enzymes are hydrolases, particularly lipases and proteases. Papain, a cysteine protease from papaya, is the best-known plant-derived hydrolytic enzyme. Bromelain from pineapple and ficain from fig latex are similar cysteine proteases that have also found applications in biocatalysis. Horseradish peroxidase is a versatile oxidative enzyme obtained from its namesake that oxidizes many organic compounds, especially phenols. Hydroxynitrile lyase (often called oxynitrilase) from bitter almond is one of the most important plant-derived... 

In enzymes, the most common nucleophilic groups that are functional in catalysis are the serine hydroxyl—which occurs in the serine proteases, cholinesterases, esterases, lipases, and alkaline phosphatases—and the cysteine thiol—which occurs in the thiol proteases (papain, ficin, and bromelain), in glyceraldehyde 3-phosphate dehydrogenase, etc. The imidazole of histidine usually functions as an acid-base catalyst and enhances the nucleophilicity of hydroxyl and thiol groups, but it sometimes acts as a nucleophile with the phos-phoryl group in phosphate transfer (Table 2.5). 

Cysteine (or thiol) proteases. The /3-thiol group is an even better nucleophile in comparison with the serine side chain thiol proteases operate under alkaline pH values as well examples include papain and bromelain. 

Trp, Leu, Met), and it was selected for making the Tyr-Gly, Phe-Leu, and Phe-Met peptide bonds. Papain was selected for Gly-Gly and Gly-Phe peptide bonds. Bromelain is very similar to papain as far as its specificity. All these proteases are serine or cysteine type, and the peptide bond formation can be done under kinetically controlled conditions. Thermolysin is an asparyl protease, and it was selected mainly for Phe-Leu and Gly-Phe bonds. MeCN, EtOAc, and methyl caproate were used as solvents with a controlled amount of buffer or at fixed water amount. 

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