Down-regulation of human extracellular cysteine protease inhibitors by the secreted staphylococcal cysteine proteases, staphopain A and B

2007 ◽  
Vol 388 (4) ◽  
pp. 437-446 ◽  
Author(s):  
Bjarne Vincents ◽  
Patrik Önnerfjord ◽  
Milosz Gruca ◽  
Jan Potempa ◽  
Magnus Abrahamson
Keyword(s):  
Cystatin C ◽  
Cathepsin B ◽  
Time Course ◽  
Cysteine Proteases ◽  
Normal Activity ◽  
Therapeutic Agents ◽  
Cysteine Protease Inhibitors ◽  
Protein Substrates ◽  
Cathepsin H

Abstract Of seven human cystatins investigated, none inhibited the cysteine proteases staphopain A and B secreted by the human pathogen Staphylococcus aureus. Rather, the extracellular cystatins C, D and E/M were hydrolyzed by both staphopains. Based on MALDI-TOF time-course experiments, staphopain A cleavage of cystatin C and D should be physiologically relevant and occur upon S. aureus infection. Staphopain A hydrolyzed the Gly11 bond of cystatin C and the Ala10 bond of cystatin D with similar K m values of approximately 33 and 32 μM, respectively. Such N-terminal truncation of cystatin C caused >300-fold lower inhibition of papain, cathepsin B, L and K, whereas the cathepsin H activity was compromised by a factor of ca. 10. Similarly, truncation of cystatin D caused alleviated inhibition of all endogenous target enzymes investigated. The normal activity of the cystatins is thus down-regulated, indicating that the bacterial enzymes can cause disturbance of the host protease-inhibitor balance. To illustrate the in vivo consequences, a mixed cystatin C assay showed release of cathepsin B activity in the presence of staphopain A. Results presented for the specificity of staphopains when interacting with cystatins as natural protein substrates could aid in the development of therapeutic agents directed toward these proteolytic virulence factors.

scholarly journals Differential Localization of Cysteine Protease Inhibitors and a Target Cysteine Protease, Cathepsin B, by Immuno-Confocal Microscopy

1998 ◽  
Vol 46 (6) ◽  
pp. 745-751 ◽  
Author(s):  
Cathárine C. Calkins ◽  
Mansoureh Sameni ◽  
Jennifer Koblinski ◽  
Bonnie F. Sloane ◽  
Kamiar Moin
Keyword(s):  
Confocal Microscopy ◽  
Cell Line ◽  
Protease Inhibitors ◽  
Cystatin C ◽  
Cysteine Protease ◽  
Cathepsin B ◽  
Cysteine Proteases ◽  
Hepatoma Cells ◽  
Embryonic Liver ◽  
Cysteine Protease Inhibitors

The cystatin superfamily of cysteine protease inhibitors and target cysteine proteases such as cathepsin B have been implicated in malignant progression. The respective cellular/extracellular localization of cystatins and cysteine proteases in tumors may be critical in regulating activity of the enzymes. Confocal microscopy has enabled us to demonstrate the differential localization of cystatins and cathepsin B in an embryonic liver cell line and an invasive hepatoma cell line. In both, stefins A and B were distributed diffusely throughout the cytoplasm, whereas cystatin C was distributed in juxtanuclear vesicles. Stefin A and cystatin C, but not stefin B, were present on the cell surface. Cystatin C was found on the top surfaces of both cell lines, whereas stefin A was found only on the top surface of the embryonic liver cells. Cathepsin B staining was concentrated in perinuclear vesicles in the embryonic liver cells. In the hepatoma cells, staining for cathepsin B was also present in vesicles adjacent to the cell membrane and on localized regions of the bottom surface. Such a disparate distribution of cathepsin B and its endogenous inhibitors may facilitate proteolysis by the hepatoma cells and thereby contribute to their invasive phenotype.

Cathepsin B, Cathepsin H, Cathepsin X and Cystatin C in Sera of Patients with Early-Stage and Inflammatory Breast Cancer

2008 ◽  
Vol 23 (3) ◽  
pp. 161-168 ◽  
Author(s):  
J. Decock ◽  
N. Obermajer ◽  
S. Vozelj ◽  
W. Hendrickx ◽  
R. Paridaens ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cystatin C ◽  
Inflammatory Breast Cancer ◽  
Tumor Size ◽  
Cathepsin B ◽  
Early Stage ◽  
Menopausal Status ◽  
Preoperative Serum ◽  
Cathepsin H ◽  
Cathepsin X

Numerous studies have linked cathepsins and their inhibitor cystatin C to tumor invasion and metastasis. We examined whether cathepsin B, cathepsin H, cathepsin X and cystatin C could be detected in sera from women with early-stage or inflammatory breast cancer and whether they correlated with clinicopathological characteristics. Preoperative serum was obtained from 176 patients with early-stage breast cancer (tumor size <5 cm, negative lymph nodes) and 31 patients with inflammatory breast cancer. Cathepsin and cystatin C levels were measured by ELISA. The patient and tumor characteristics under study were age at diagnosis, menopausal status, tumor size, tumor grade, and steroid hormone receptor status. Serum cathepsin B levels were significantly lower in patients with poorly differentiated tumors. High cystatin C levels were associated with tumor size, postmenopausal status and patient age. Interestingly, significantly lower levels of cathepsin X and H were found in patients with inflammatory breast cancer, a trend also observed for cathepsin B and cystatin C. In conclusion, our results show a limited association of cathepsins B, H, X and cystatin C with established prognostic parameters. These data are promising and encourage future analysis of the clinical outcome of our patients in order to examine the potential prognostic value of these biomarkers. Further, this study indicates a role for cathepsin X and H in inflammatory breast cancer.

scholarly journals Structure-Activity Relationships for Inhibition of Cysteine Protease Activity and Development of Plasmodium falciparum by Peptidyl Vinyl Sulfones

2003 ◽  
Vol 47 (1) ◽  
pp. 154-160 ◽  
Author(s):  
Bhaskar R. Shenai ◽  
Belinda J. Lee ◽  
Alejandro Alvarez-Hernandez ◽  
Pek Y. Chong ◽  
Cory D. Emal ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Cysteine Protease ◽  
Cysteine Proteases ◽  
Parasite Development ◽  
Vinyl Sulfone ◽  
Cysteine Protease Inhibitors ◽  
Structure Activity Relationships ◽  
Structure Activity ◽  
Sulfonate Ester

ABSTRACT The Plasmodium falciparum cysteine proteases falcipain-2 and falcipain-3 appear to be required for hemoglobin hydrolysis by intraerythrocytic malaria parasites. Previous studies showed that peptidyl vinyl sulfone inhibitors of falcipain-2 blocked the development of P. falciparum in culture and exerted antimalarial effects in vivo. We now report the structure-activity relationships for inhibition of falcipain-2, falcipain-3, and parasite development by 39 new vinyl sulfone, vinyl sulfonate ester, and vinyl sulfonamide cysteine protease inhibitors. Levels of inhibition of falcipain-2 and falcipain-3 were generally similar, and many potent compounds were identified. Optimal antimalarial compounds, which inhibited P. falciparum development at low nanomolar concentrations, were phenyl vinyl sulfones, vinyl sulfonate esters, and vinyl sulfonamides with P2 leucine moieties. Our results identify independent structural correlates of falcipain inhibition and antiparasitic activity and suggest that peptidyl vinyl sulfones have promise as antimalarial agents.

scholarly journals Molecular and Biochemical Characterization of a Cathepsin B-Like Protease Family Unique to Trypanosoma congolense

2008 ◽  
Vol 7 (4) ◽  
pp. 684-697 ◽  
Author(s):  
Carlos Mendoza-Palomares ◽  
Nicolas Biteau ◽  
Christiane Giroud ◽  
Virginie Coustou ◽  
Theresa Coetzer ◽  
...  
Keyword(s):  
Cathepsin B ◽  
Biochemical Characterization ◽  
Catalytic Domain ◽  
Expression Profiles ◽  
Specific Inhibitor ◽  
Cysteine Proteases ◽  
Catalytic Triad ◽  
Trypanosoma Congolense

ABSTRACT Cysteine proteases have been shown to be essential virulence factors and drug targets in trypanosomatids and an attractive antidisease vaccine candidate for Trypanosoma congolense. Here, we describe an important amplification of genes encoding cathepsin B-like proteases unique to T. congolense. More than 13 different genes were identified, whereas only one or two highly homologous genes have been identified in other trypanosomatids. These proteases grouped into three evolutionary clusters: TcoCBc1 to TcoCBc5 and TcoCBc6, which possess the classical catalytic triad (Cys, His, and Asn), and TcoCBs7 to TcoCBs13, which contains an unusual catalytic site (Ser, Xaa, and Asn). Expression profiles showed that members of the TcoCBc1 to TcoCBc5 and the TcoCBs7 to TcoCBs13 groups are expressed mainly in bloodstream forms and localize in the lysosomal compartment. The expression of recombinant representatives of each group (TcoCB1, TcoCB6, and TcoCB12) as proenzymes showed that TcoCBc1 and TcoCBc6 are able to autocatalyze their maturation 21 and 31 residues, respectively, upstream of the predicted start of the catalytic domain. Both displayed a carboxydipeptidase function, while only TcoCBc1 behaved as an endopeptidase. TcoCBc1 exhibited biochemical differences regarding inhibitor sensitivity compared to that of other cathepsin B-like proteases. Recombinant pro-TcoCBs12 did not automature in vitro, and the pepsin-matured enzyme was inactive in tests with cathepsin B fluorogenic substrates. In vivo inhibition studies using CA074Me (a cell-permeable cathepsin B-specific inhibitor) demonstrated that TcoCB are involved in lysosomal protein degradation essential for survival in bloodstream form. Furthermore, TcoCBc1 elicited an important immune response in experimentally infected cattle. We propose this family of proteins as a potential therapeutic target and as a plausible antigen for T. congolense diagnosis.

scholarly journals The Homology Modeling and Docking Investigation of Human Cathepsin B

2020 ◽  
Vol 10 (1) ◽  
pp. 26687
Author(s):  
Afshin Khara ◽  
Ehsan Jahangirian ◽  
Hossein Tarrahimofrad
Keyword(s):  
Amino Acids ◽  
Protease Inhibitors ◽  
Cystatin C ◽  
Active Site ◽  
Cathepsin B ◽  
Antigen Processing ◽  
3D Structure ◽  
Cysteine Proteases ◽  
Docking Studies ◽  
Alternative Methods

Background: Cathepsin B comprises a group of lysosomal cysteine proteases belonging to the Papain family; it has an intracellular function in the process of protein catabolism, antigen processing in the immune response, and Alzheimer’s disease. In cancers, cathepsin B interferes with autophagy and intracellular catabolism, and breaks down extracellular matrix, decreases protease inhibitors expression, and ultimately helps to accelerate metastasis, tumor malignancy, and reduce immune resistance. Methods: In this study, the 3D structure of cathepsin B was constructed using modeler and Iterative Threading ASSEmbly Refinement (I-TASSER), based on similarity to the crystallographic model of procathepsin B (1PBH). Then, the predicted cathepsin B model was evaluated using PROCHECK and PROSA for quality and reliability. Molecular studies suggested that the amino acids cysteine 108, histidine 189, and histidine 190 form the envelope of the active site of cathepsin B. The docking studies of cathepsin B was performed with protease inhibitors cystatin C, E-64 and leupeptin. Results: The lowest binding energy was related to the cathepsin B-E-64 complex. Accordingly, it was found that E64 interacts with the amino acid cysteine 108 of the active site of cathepsin B. Leupeptin made 2 hydrogen bonds with cathepsin B, but none with the active site of cathepsin amino acids. Cystatin C established a hydrogen bond with the arginine 18 of cathepsin B and made electrostatic bonds with aspartate 148 of cathepsin B. Conclusion: Therefore, the bioinformatics and docking studies of cathepsin B with its inhibitors could be used as reliable identification, treatment, and alternative methods for selecting the inhibitors and controllers of cancer progression.

scholarly journals Cathepsin Cs Are Key for the Intracellular Survival of the Protozoan Parasite, Toxoplasma gondii

2006 ◽  
Vol 282 (7) ◽  
pp. 4994-5003 ◽  
Author(s):  
Xuchu Que ◽  
Juan C. Engel ◽  
David Ferguson ◽  
Annette Wunderlich ◽  
Stanislas Tomavo ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Cathepsin B ◽  
Parasitophorous Vacuole ◽  
Cathepsin L ◽  
Protozoan Parasite ◽  
Cysteine Proteases ◽  
Intracellular Survival ◽  
Chemotherapeutic Agents ◽  
Cathepsin C

Cysteine proteases play key roles in apicomplexan invasion, organellar biogenesis, and intracellular survival. We have now characterized five genes encoding papain family cathepsins from Toxoplasma gondii, including three cathepsin Cs, one cathepsin B, and one cathepsin L. Unlike endopeptidases cathepsin B and L, T. gondii cathepsin Cs are exopeptidases and remove dipeptides from unblocked N-terminal substrates of proteins or peptides. TgCPC1 was the most highly expressed cathepsin mRNA in tachyzoites (by real-time PCR), but three cathepsins, TgCPC1, TgCPC2, and TgCPB, were undetectable in in vivo bradyzoites. The specific cathepsin C inhibitor, Gly-Phe-dimethylketone, selectively inhibited the TgCPCs activity, reducing parasite intracellular growth and proliferation. The targeted disruption of TgCPC1 does not affect the invasion and growth of tachyzoites as TgCPC2 is then up-regulated and may substitute for TgCPC1. TgCPC1 and TgCPC2 localize to constitutive secretory vesicles of tachyzoites, the dense granules. T. gondii cathepsin Cs are required for peptide degradation in the parasitophorous vacuole as the degradation of the marker protein, Escherichia coli β-lactamase, secreted into the parasitophorous vacuole of transgenic tachyzoites was completely inhibited by the cathepsin C inhibitor. Cathepsin C inhibitors also limited the in vivo infection of T. gondii in the chick embryo model of toxoplasmosis. Thus, cathepsin Cs are critical to T. gondii growth and differentiation, and their unique specificities could be exploited to develop novel chemotherapeutic agents.

scholarly journals Effect of L-arginine and L-name on lysosomal cysteine proteases activity and lysosomal membranes permeability in rat aorta

2016 ◽  
Vol 97 (2) ◽  
pp. 250-255 ◽  
Author(s):  
A I Arapova ◽  
M A Fomina
Keyword(s):  
Acid Phosphatase ◽  
Cathepsin B ◽  
Cysteine Proteases ◽  
Total Activity ◽  
Rat Aorta ◽  
Lysosomal Fraction ◽  
Lysosomal Membranes ◽  
Cathepsin H ◽  
Male Wistar Rats ◽  
Combined Drugs

Aim. To study the effect of L-arginine and its analogue N-nitro-L-arginine methyl ester (L-NAME) alone and in combination on lysosomal cysteine proteolysis and lysosomal membranes state in rat aorta.Methods. The study was performed on male Wistar rats kept under standard vivarium conditions and divided into three control and three experimental groups of 8 animals each. The experimental samples included groups with L-arginine and/or L-NAME administration. The indicators were studied in the rat aorta homogenate precipitating and non precipitating fractions. Acid phosphatase activity was determined by a standardized method of «end point», the cathepsins B, L and H activity was studied by spectrofluorimetric method.Results. When simulating the changes of nitric oxide synthesis level using L-arginine, the increase of the total cathepsins activity was detected, acid phosphatase lability coefficient was reduced, what is characterized by general lysosomal membranes stabilization. L-NAME group, in contrast, is characterized by a decrease in the cathepsin B and H activity indicators, differences from arginine group were observed in the cathepsin H in lysosomal and general fractions, lysosomal membrane is labile. Combined drugs administration reduces the total cathepsins activity, while there is an increase of the acid phosphatase total activity, all indicators suggest lysosomal membranes labilization.Conclusion. L-arginine at a dose of 500 mg/kg causes increase in the total cathepsins B, L and H activity in rat aorta due to lysosomal fraction; L-arginine action leads to lysosomal membranes stabilization; L-NAME group in cathepsin H shows a decrease in the cathepsins secretion level with decreased total activity due to both factions; combined administration of arginine + L-NAME group in cathepsin B is characterized by an increase in secretion due to lysosomes membrane labilization.

scholarly journals Defense against own arms: staphylococcal cysteine proteases and their inhibitors.

2003 ◽  
Vol 50 (3) ◽  
pp. 715-724 ◽  
Author(s):  
Grzegorz Dubin
Keyword(s):  
Staphylococcus Aureus ◽  
Protease Inhibitors ◽  
Current Knowledge ◽  
Cysteine Proteases ◽  
Infection Process ◽  
Secreted Proteins ◽  
Human Pathogen ◽  
Cysteine Protease Inhibitors ◽  
Wide Range

Staphylococcus aureus is a human pathogen causing a wide range of diseases. Most staphylococcal infections, unlike those caused by other bacteria are not toxigenic and very little is known about their pathogenesis. It has been proposed that a core of secreted proteins common to many infectious strains is responsible for colonization and infection. Among those proteins several proteases are present and over the years many different functions in the infection process have been attributed to them. However, little direct, in vivo data has been presented. Two cysteine proteases, staphopain A (ScpA) and staphopain B (SspB) are important members of this group of enzymes. Recently, two cysteine protease inhibitors, staphostatin A and staphostatin B (ScpB and SspC, respectively) were described in S. aureus shedding new light on the complexity of the processes involving the two proteases. The scope of this review is to summarize current knowledge on the network of staphylococcal cysteine proteases and their inhibitors in view of their possible role as virulence factors.

scholarly journals Targeting TMPRSS2 and Cathepsin B/L Together May Be Synergistic Against SARS-CoV-2 Infection

2020 ◽  
Author(s):  
Pranesh Padmanabhan ◽  
Rajat Desikan ◽  
Narendra Dixit
Keyword(s):  
Cathepsin B ◽  
Protein S ◽  
Cysteine Proteases ◽  
Target Cells ◽  
Dependent Manner ◽  
Dose Dependent ◽  
Protease Expression ◽  
Entry Efficiency

<p>The entry of SARS-CoV-2 into target cells requires the activation of its surface spike protein, S, by host proteases. The host serine protease TMPRSS2 and cysteine proteases Cathepsin B/L can activate S, making two independent entry pathways accessible to SARS-CoV-2. Blocking the proteases prevents SARS-CoV-2 entry <i>in vitro</i>. This blockade may be achieved <i>in vivo</i> through ‘repurposing’ drugs, a potential treatment option for COVID-19 that is now in clinical trials. Here, we found, surprisingly, that drugs targeting the two pathways, although independent, could display strong synergy in blocking virus entry. We predicted this synergy first using a mathematical model of SARS-CoV-2 entry and dynamics <i>in vitro</i>. The model considered the two pathways explicitly, let the entry efficiency through a pathway depend on the corresponding protease expression level, which varied across cells, and let inhibitors compromise the efficiency in a dose-dependent manner. The synergy predicted was novel and arose from effects of the drugs at both the single cell and the cell population levels. Validating our predictions, available <i>in vitro</i> data on SARS-CoV-2 and SARS-CoV entry displayed this synergy. Further, analysing the data using our model, we estimated the relative usage of the two pathways and found it to vary widely across cell lines, suggesting that targeting both pathways <i>in vivo</i> may be important and synergistic given the broad tissue tropism of SARS-CoV-2. Our findings provide insights into SARS-CoV-2 entry into target cells and may help improve the deployability of drug combinations targeting host proteases required for the entry. <br></p>

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