Crystal structure of cathepsin B inhibited with CA030 at 2.0-.ANG. resolution: A basis for the design of specific epoxysuccinyl inhibitors

Turk, Dušan; Podobnik, Marjetka; Popović, Tatjana; Katunuma, Nobuhiko; Bode, Wolfram; Huber, Robert; Türk, Vito

doi:10.1021/bi00014a037

Cited by 184 publications

(200 citation statements)

References 25 publications

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“…It may be because the peptide is oriented in the reverse position compared to a natural peptide substrate. This reverse orientation has been described for the interaction between cathepsin B and its proregion in the rat procathepsin B crystal [13], and it also corresponds to the orientation of E 64 and its derivatives with papain and cathepsin B [25][26][27][28]. This explanation is supported by the observation that the same peptide with an alkylated cysteinyl residue (PB11Acre) is more rapidly and unspecifically degraded by cathepsin B. Peptide PBll therefore contains cleavage sites, but they are not accessible.…”

Section: Resultsmentioning

confidence: 57%

Inhibition of cathepsin B by its propeptide: Use of overlapping peptides to identify a critical segment

et al. 1996

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Ten overlapping 15-mer peptides (peptidyl amides) spanning the proregion of rat cathepsin B (residues lp-60p) were constructed to identify minimal segments having inhibitory activity towards the mature enzyme, that could be used to develop a new generation of peptide-derived inhibitors specifically targeting the active site of the corresponding proteinase. Three synthetic peptides, containing the pentapeptide Leu-CysGly-Thr-Val (residues 41p-45p) in their sequence, inhibited cathepsin B with Ki values in the micromolar range. Alkylation of the thiol group of Cys-42p of peptide PB8 (36p-50p) resulted in its rapid proteolytic degradation, suggesting that this residue is essential for inhibition. The inhibition constant was slightly improved (Kt = 2 ltM) using a longer peptide (26p-50p) which was completely resistant to cleavage even after a prolonged incubation. Alkylation of its cysteinyl residue also resulted in rapid cleavage of the peptide chain. Peptides derived from the rat cathepsin B prosequence also inhibited human cathepsin B with similar Kl values. Unlike rat cathepsin B, which cleaves peptide PB8 at the G47p-G48p bond after prolonged incubation, the human enzyme cleaved both PB8 and PBll at the Lys-40p-Leu41p bond, in agreement with the different kinetic properties of these two proteinases. New probes with improved specificity for cysteine proteinases may therefore be designed based on the sequences of their propeptides.

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Section: Resultsmentioning

confidence: 57%

Inhibition of cathepsin B by its propeptide: Use of overlapping peptides to identify a critical segment

et al. 1996

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“…The crystal structures of human and rat cathepsin B [13,14] have confirmed high structural similarity to plant enzyme papain [16] and structural reasons for cathepsin B peptidyl dipeptidase activity [13,15] have also been revealed. An understanding of the molecular basis for the activation mechanism [17 19] of cathepsin B requires knowledge of the 3-dimensional structure of the proenzyme.…”

Section: Introductionmentioning

confidence: 71%

Crystal structures of human procathepsin B at 3.2 and 3.3 Å resolution reveal an interaction motif between a papain‐like cysteine protease and its propeptide

et al. 1996

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A wild-type human procathepsin B was expressed, crystallized in two crystal° forms and its crystal structure determined at 3.2 and 3.3 A resolution. The structure reveals that the propeptide folds on the cathepsin B surface, shielding the enzyme active site from exposure to solvent. The structure of the enzymaticaily active domains is virtually identical to that of the native enzyme [Musil et al. (1991) EMBO J. 10, 2321-2330]: the main difference is that the occluding loop residues are lifted above the body of the mature enzyme, supporting the propeptide structure.

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“…These structural features are illustrated with a crystal structure of papain (1ppp; Kim et al, 1992), which belongs to XCP2-like PLCPs (subfamily 3) and represents subfamilies 1 to 8. CTB3-like proteases (subfamily 9) are illustrated with a crystal structure of human cathepsin B (4csb; Turk et al, 1995;Fig. 3A).…”

Section: Structural Features Are Subfamily Specificmentioning

confidence: 99%

Subclassification and Biochemical Analysis of Plant Papain-Like Cysteine Proteases Displays Subfamily-Specific Characteristics

Richau

Kaschani

Verdoes³

et al. 2012

Plant Physiology

169

222

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Papain-like cysteine proteases (PLCPs) are a large class of proteolytic enzymes associated with development, immunity, and senescence. Although many properties have been described for individual proteases, the distribution of these characteristics has not been studied collectively. Here, we analyzed 723 plant PLCPs and classify them into nine subfamilies that are present throughout the plant kingdom. Analysis of these subfamilies revealed previously unreported distinct subfamily-specific functional and structural characteristics. For example, the NPIR and KDEL localization signals are distinctive for subfamilies, and the carboxyl-terminal granulin domain occurs in two PLCP subfamilies, in which some individual members probably evolved by deletion of the granulin domains. We also discovered a conserved double cysteine in the catalytic site of SAG12-like proteases and two subfamily-specific disulfides in RD19A-like proteases. Protease activity profiling of representatives of the PLCP subfamilies using novel fluorescent probes revealed striking polymorphic labeling profiles and remarkably distinct pH dependency. Competition assays with peptide-epoxide scanning libraries revealed common and unique inhibitory fingerprints. Finally, we expand the detection of PLCPs by identifying common and organ-specific protease activities and identify previously undetected proteases upon labeling with cell-penetrating probes in vivo. This study provides the plant protease research community with tools for further functional annotation of plant PLCPs.

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Crystal structure of cathepsin B inhibited with CA030 at 2.0-.ANG. resolution: A basis for the design of specific epoxysuccinyl inhibitors

Cited by 184 publications

References 25 publications

Inhibition of cathepsin B by its propeptide: Use of overlapping peptides to identify a critical segment

Inhibition of cathepsin B by its propeptide: Use of overlapping peptides to identify a critical segment

Crystal structures of human procathepsin B at 3.2 and 3.3 Å resolution reveal an interaction motif between a papain‐like cysteine protease and its propeptide

Subclassification and Biochemical Analysis of Plant Papain-Like Cysteine Proteases Displays Subfamily-Specific Characteristics

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