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Papain Family (papain + family)

Distribution by Scientific Domains

Chemistry	100%

Selected Abstracts

Subsite specificity of trypanosomal cathepsin L-like cysteine proteases

FEBS JOURNAL, Issue 9 2001
Probing the S2 pocket with phenylalanine-derived amino acids
The S2 subsite of mammalian cysteine proteinases of the papain family is essential for specificity. Among natural amino acids, all these enzymes prefer bulky hydrophobic residues such as phenylalanine at P2. This holds true for their trypanosomal counterparts: cruzain from Trypanosoma cruzi and congopain from T. congolense. A detailed analysis of the S2 specificity of parasitic proteases was performed to gain information that might be of interest for the design of more selective pseudopeptidyl inhibitors. Nonproteogenic phenylalanyl analogs (Xaa) have been introduced into position P2 of fluorogenic substrates dansyl-Xaa-Arg-Ala-Pro-Trp, and their kinetic constants (Km, kcat/Km) have been determined with congopain and cruzain, and related host cathepsins B and L. Trypanosomal cysteine proteases are poorly stereoselective towards d/l -Phe, the inversion of chirality modifying the efficiency of the reaction but not the Km. Congopain binds cyclohexylalanine better than aromatic Phe derivatives. Another characteristic feature of congopain compared to cruzain and cathepsins B and L was that it could accomodate a phenylglycyl residue (kcat/Km = 1300 mm,1·s,1), while lengthening of the side chain by a methylene group only slightly impaired the specificity constant towards trypanosomal cysteine proteases. Mono- and di-halogenation or nitration of Phe did not affect Km for cathepsin L-like enzymes, but the presence of constrained Phe derivatives prevented a correct fitting into the S2 subsite. A model of congopain has been built to study the fit of Phe analogs within the S2 pocket. Phe analogs adopted a positioning within the S2 pocket similar to that of the Tyr of the cruzain/Z-Tyr-Ala-fluoromethylketone complex. However, cyclohexylalanine has an energetically favorable chair-like conformation and can penetrate deeper into the subsite. Fitting of modeled Phe analogs were in good agreement with kinetic parameters. Furthermore, a linear relationship could be established with logP, supporting the suggestion that fitting into the S2 pocket of trypanosomal cysteine proteases depends on the hydrophobicity of Phe analogs. [source]

Potency and selectivity of inhibition of cathepsin K, L and S by their respective propeptides

FEBS JOURNAL, Issue 20 2000
Jocelyne Guay
The prodomains of several cysteine proteases of the papain family have been shown to be potent inhibitors of their parent enzymes. An increased interest in cysteine proteases inhibitors has been generated with potential therapeutic targets such as cathepsin K for osteoporosis and cathepsin S for immune modulation. The propeptides of cathepsin S, L and K were expressed as glutathione S -transferase-fusion proteins in Escherichia coli. The proteins were purified on glutathione affinity columns and the glutathione S -transferase was removed by thrombin cleavage. All three propeptides were tested for inhibitor potency and found to be selective within the cathepsin L subfamily (cathepsins K, L and S) compared with cathepsin B or papain. Inhibition of cathepsin K by either procathepsin K, L or S was time-dependent and occurred by an apparent one-step mechanism. The cathepsin K propeptide had a Ki of 3.6,6.3 nm for each of the three cathepsins K, L and S. The cathepsin L propeptide was at least a 240-fold selective inhibitor of cathepsin K (Ki = 0.27 nm) and cathepsin L (Ki = 0.12 nm) compared with cathepsin S (Ki = 65 nm). Interestingly, the cathepsin S propeptide was more selective for inhibition of cathepsin L (Ki = 0.46 nm) than cathepsin S (Ki = 7.6 nm) itself or cathepsin K (Ki = 7.0 nm). This is in sharp contrast to previously published data demonstrating that the cathepsin S propeptide is equipotent for inhibition of human cathepsin S and rat and paramecium cathepsin L [Maubach, G., Schilling, K., Rommerskirch, W., Wenz, I., Schultz, J.E., Weber, E. & Wiederanders, B. (1997), Eur J. Biochem. 250, 745,750]. These results demonstrate that limited selectivity of inhibition can be measured for the procathepsins K, L and S vs. the parent enzymes, but selective inhibition vs. cathepsin B and papain was obtained. [source]

Structure of the mexicain,E-64 complex and comparison with other cysteine proteases of the papain family

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 5 2007
J. A. Gavira
Mexicain is a 23.8,kDa cysteine protease from the tropical plant Jacaratia mexicana. It is isolated as the most abundant product after cation-exchange chromatography of the mix of proteases extracted from the latex of the fruit. The purified enzyme inhibited with E-64 [N -(3-carboxyoxirane-2-carbonyl)-leucyl-amino(4-guanido)butane] was crystallized by sitting-drop vapour diffusion and the structure was solved by molecular replacement at 2.1,Å resolution and refined to an R factor of 17.7% (Rfree = 23.8%). The enzyme belongs to the ,+, class of proteins and the structure shows the typical papain-like fold composed of two domains, the ,-helix-rich (L) domain and the ,-barrel-like (R) domain, separated by a groove containing the active site formed by residues Cys25 and His159, one from each domain. The four monomers in the asymmetric unit show one E-64 molecule covalently bound to Cys25 in the active site and differences have been found in the placement of E-64 in each monomer. [source]

The ribbon of hydrogen bonds in globular proteins.

BIOPOLYMERS, Issue 2 2004

Abstract A study of the role of the hydrogen-bonding side chains in the ribbon of hydrogen bonds in globular proteins, using the papain family as an example, suggests that these side chains may be divided into three categories depending on their position in the molecule. In the first category, they form part of the local ribbon, in the second they form part of the ribbon at a site remote along the main chain, and in the third they play no role in the formation of the ribbon. The second case is particularly interesting because it provides a natural mechanism for the formation of the tertiary structure of the globular proteins. The results suggest that the robustness of the globular proteins towards mutations arises from the fact that many mutations that involve hydrogen-bonding side chains either leave the hydrogen bonding of the ribbon essentially unchanged or their hydrogen bonding plays no part in the formation of the ribbon in the first place. The results show that it is possible to obtain the ribbon of hydrogen bonds for a family of proteins whose data set's are of intermediate quality by studying the ribbons of several members of such a family and then taking an average over the different partial ribbons to create a standard ribbon of hydrogen bonds for the family as a whole. This method is used here to derive the standard ribbon for the papain family with papain itself, actinidin, and human liver cathepsin B as the representatives of the family. All three members of the family fit the standard ribbon with an accuracy of 85,91%. This result opens up the use of this technique for the study of a large number of globular proteins whose recorded data sets are of intermediate quality. © 2003 Wiley Periodicals, Inc. Biopolymers 73: 178,191, 2004 [source]