Characterisation of a highly specific, endogenous inhibitor of cysteine protease from Staphylococcus epidermidis, a new member of the staphostatin family

2004 ◽  
Vol 385 (6) ◽  
pp. 543-546 ◽  
Author(s):  
G. Dubin ◽  
J. Stec-Niemczyk ◽  
T. Dylag ◽  
J. Silberring ◽  
A. Dubin ◽  
...  
Keyword(s):  
Cysteine Protease ◽  
Staphylococcus Epidermidis ◽  
Peptide Bond ◽  
Cysteine Proteases ◽  
Proteolytic Cleavage ◽  
Reactive Site ◽  
Cysteine Protease Inhibitors ◽  
The Family ◽  
Mechanism Of Interaction ◽  
The Difference

AbstractStaphostatins, a novel family of cysteine protease inhibitors with a unique mechanism of action and distinct protein fold has recently been discovered. In this report we describe the properties ofStaphylococcus epidermidisstaphostatin A (EcpB), a new member of the family. As for other staphostatins, the recombinantS. epidermidisstaphostatin A exerted very narrow inhibitory specificity, limited to cysteine protease from the same species. The closely related proteases fromS. aureuscleaved the inhibitor at the reactive site peptide bond and inactivated it. The EcpB homologue,S. aureusstaphostatin A (ScpB), was also susceptible to proteolytic cleavage at the same site by nontarget cysteine proteases. Conversely,S. aureusstaphostatin B (SspC) was resistant to such proteolysis. The difference in the susceptibility of individual inhibitors to proteolytic cleavage at the reactive site suggests subtle variations in the mechanism of interaction with cysteine proteases.

scholarly journals Inhibition of Staphylococcus aureus cysteine proteases by human serpin potentially limits staphylococcal virulence

2011 ◽  
Vol 392 (5) ◽  
Author(s):  
Tomasz Kantyka ◽  
Karolina Plaza ◽  
Joanna Koziel ◽  
Danuta Florczyk ◽  
Hennig R. Stennicke ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Serine Proteases ◽  
Peptide Bond ◽  
Cysteine Proteases ◽  
Reactive Site ◽  
Association Rate ◽  
High Association ◽  
Endogenous Protease ◽  
Covalent Complex

AbstractBacterial proteases are considered virulence factors and it is presumed that by abrogating their activity, host endogenous protease inhibitors play a role in host defense against invading pathogens. Here we present data showing thatStaphylococcus aureuscysteine proteases (staphopains) are efficiently inhibited by Squamous Cell Carcinoma Antigen 1 (SCCA1), an epithelial-derived serpin. The high association rate constant (kass) for inhibitory complex formation (1.9×104m/s and 5.8×104 m/s for staphopain A and staphopain B interaction with SCCA1, respectively), strongly suggests that SCCA1 can regulate staphopain activityin vivoat epithelial surfaces infected/colonized byS. aureus. The mechanism of staphopain inhibition by SCCA1 is apparently the same for serpin interaction with target serine proteases whereby the formation of a covalent complex result in cleavage of the inhibitory reactive site peptide bond and associated release of the C-terminal serpin fragment. Interestingly, the SCCA1 reactive site closely resembles a motif in the reactive site loop of nativeS. aureus-derived inhibitors of the staphopains (staphostatins). Given thatS. aureusis a major pathogen of epithelial surfaces, we suggest that SCCA1 functions to temper the virulence of this bacterium by inhibiting the staphopains.

scholarly journals Structure-Function of Falcipains: Malarial Cysteine Proteases

2012 ◽  
Vol 2012 ◽  
pp. 1-11 ◽  
Author(s):  
Kailash C. Pandey ◽  
Rajnikant Dixit
Keyword(s):  
Functional Analysis ◽  
Drug Development ◽  
Protease Inhibitors ◽  
Cysteine Protease ◽  
Essential Role ◽  
Cysteine Proteases ◽  
Structural Studies ◽  
Cysteine Protease Inhibitors ◽  
Hemoglobin Binding ◽  
Structural Insight

Evidence indicates that cysteine proteases play essential role in malaria parasites; therefore an obvious area of investigation is the inhibition of these enzymes to treat malaria. Studies with cysteine protease inhibitors and manipulating cysteine proteases genes have suggested a role for cysteine proteases in hemoglobin hydrolysis. The best characterizedPlasmodiumcysteine proteases are falcipains, which are papain family enzymes. Falcipain-2 and falcipain-3 are major hemoglobinases ofP. falciparum. Structural and functional analysis of falcipains showed that they have unique domains including a refolding domain and a hemoglobin binding domain. Overall, the complexes of falcipain-2 and falcipain-3 with small and macromolecular inhibitors provide structural insight to facilitate the design or modification of effective drug treatment against malaria. Drug development targeting falcipains should be aided by a strong foundation of biochemical and structural studies.

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.

Synthesis of quinoline derivatives as potential cysteine protease inhibitors

2020 ◽  
Author(s):  
Marina MS Andrade ◽  
Luan C Martins ◽  
Gabriel VL Marques ◽  
Carla A Silva ◽  
Gilson Faria ◽  
...  
Keyword(s):  
Trypanosoma Cruzi ◽  
Protease Inhibitors ◽  
Cysteine Protease ◽  
Inhibitory Activity ◽  
Molecular Targets ◽  
Cysteine Proteases ◽  
Leishmania Species ◽  
Molecular Target ◽  
Quinoline Derivatives ◽  
Cysteine Protease Inhibitors

Aim: Cysteine proteases are important molecular targets involved in the replication, virulence and survival of parasitic organisms, including Trypanosoma and Leishmania species. Methodology & results: Analogs of the 7-chloro- N-[3-(morpholin-4-yl)propyl]quinolin-4-amine were synthesized and their inhibitory activity against the enzymes cruzain and rhodesain as well as against promastigotes forms of Leishmania species and epimastigotes forms of Trypanosoma cruzi were evaluated. Five compounds showed activity against both enzymes with IC50 values ranging from 23 to 123 μM. Among these, compounds 3 and 4 displayed leishmanicidal activity; compound 4 was the most promising with IC50 values <10 μM and no cytotoxicity for uninfected cells. Conclusion: The results obtained indicate that cysteine proteases are likely to be the molecular target of compounds 3 and 4.

scholarly journals Trypanosomatid cysteine protease activity may be enhanced by a kininogen-like moiety from host serum

1995 ◽  
Vol 305 (2) ◽  
pp. 549-556 ◽  
Author(s):  
J D Lonsdale-Eccles ◽  
G W N Mpimbaza ◽  
Z R M Nkhungulu ◽  
J Olobo ◽  
L Smith ◽  
...  
Keyword(s):  
Molecular Mass ◽  
Protease Inhibitors ◽  
Cysteine Protease ◽  
Leishmania Donovani ◽  
Leishmania Major ◽  
Cysteine Proteases ◽  
Cysteine Protease Inhibitor ◽  
Cysteine Protease Inhibitors ◽  
African Trypanosomes

African trypanosomes contain cysteine proteases (trypanopains) the activity of which can be measured by in vitro digestion of fibrinogen, after electrophoresis in fibrinogen-containing SDS/polyacrylamide gels. When assessed by this procedure, trypanopain from Trypanosoma brucei (trypanopain-Tb) is estimated to have a molecular mass of 28 kDa. However, two additional bands of trypanopain activity (87 kDa and 105 kDa) are observed if serum is added to the trypanopain before electrophoresis. Formation of the 87 and 105 kDa bands is frequently accompanied by a reduction in the intensity of the 28 kDa activity which suggests that the extra bands are complexes of the 28 kDa trypanopain-Tb and a molecule from rat serum called rat trypanopain moledulator (rTM). The rTM-induced activation of cysteine proteases is not restricted to T. brucei as it is also observed with proteases from other protozoan parasites such as bloodstream forms of Trypanosoma congolense and the mammalian-infective in vitro-derived promastigote forms of Leishmania donovani and Leishmania major. The physical properties of rTM resemble those of the kininogen family of cysteine protease inhibitors. rTM is an acidic (pI 4.7) heat-stable 68 kDa glycoprotein with 15 kDa protease-susceptible domains. This resemblance between rTM and kininogens was confirmed by the positive, albeit weak, immunoreactivity between anti-(human low-molecular-mass kininogen) antibody and rTM as well as anti-rTM antibody and human low-molecular-mass kininogen. Furthermore, commercial preparations of human-low-molecular-mass kininogen and chicken egg white cystatin mimicked rTM by forming extra bands of proteolytic activity in the presence of trypanopain-Tb. In some instances, low-molecular-mass kininogen was also observed to increase the rate of hydrolysis of 7-(benzyloxycarbonyl-phenylalanyl-arginyl-amido)-4- methylcoumarin by live T. brucei. Although this effect was rather erratic, in no instance was significant inhibition observed when this putative cysteine protease inhibitor was used under these conditions. The activation of parasite cysteine proteases by commonly accepted cysteine protease inhibitors is unexpected and may have important pathological repercussions.

scholarly journals Aziridine-2,3-Dicarboxylates, Peptidomimetic Cysteine Protease Inhibitors with Antileishmanial Activity

2006 ◽  
Vol 50 (7) ◽  
pp. 2439-2447 ◽  
Author(s):  
Alicia Ponte-Sucre ◽  
Radim Vicik ◽  
Martina Schultheis ◽  
Tanja Schirmeister ◽  
Heidrun Moll
Keyword(s):  
Protease Inhibitors ◽  
Infection Rate ◽  
Cysteine Protease ◽  
Cysteine Proteases ◽  
Host Cells ◽  
Interleukin 12 ◽  
Cysteine Protease Inhibitors ◽  
Macrophage Infection ◽  
Simultaneous Treatment ◽  
Necrosis Factor Alpha

ABSTRACT Chemotherapy of leishmaniasis is mainly based on antimonials. However, they are extremely toxic and cause serious side effects, and there is a worldwide increasing frequency of chemoresistance to antimonials. These issues emphasize the urgent need for affordable alternative drugs against leishmaniasis. Leishmania cysteine proteases are essential for parasite growth, differentiation, pathogenicity, and virulence and are thus attractive targets for combating leishmaniasis. Herein we demonstrate that the cysteine protease inhibitors aziridine-2,3-dicarboxylates 13b and 13e impaired promastigote growth at mid-micromolar concentrations and decreased the infection rate of peritoneal macrophages at concentrations 8- to 13-fold lower than those needed to inhibit parasite replication. Simultaneous treatment of infected cells with compound 13b and gamma interferon resulted in an even further reduction of the concentration needed for a significant decrease in macrophage infection rate. Notably, treatment with the compounds alone modulated the cytokine secretion of infected macrophages, with increased levels of interleukin-12 and tumor necrosis factor alpha. Furthermore, the decreased infection rate in the presence of compound 13b correlated with increased nitric oxide production by macrophages. Importantly, at the concentrations used herein, compounds 13b and 13e were not toxic against fibroblasts, macrophages, or dendritic cells. Together, these results suggest that the aziridine-2,3-dicarboxylates 13b and 13e are potential antileishmanial lead compounds with low toxicity against host cells and selective antiparasitic effects.

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.

Expression and characterization of the Plasmodium falciparum haemoglobinase falcipain-3

2001 ◽  
Vol 360 (2) ◽  
pp. 481-489 ◽  
Author(s):  
Puran S. SIJWALI ◽  
Bhaskar R. SHENAI ◽  
Jiri GUT ◽  
Ajay SINGH ◽  
Philip J. ROSENTHAL
Keyword(s):  
Plasmodium Falciparum ◽  
Protease Inhibitors ◽  
Cysteine Protease ◽  
Active Form ◽  
Cysteine Proteases ◽  
Food Vacuole ◽  
Acidic Ph ◽  
Cysteine Protease Inhibitors ◽  
Peptide Substrates ◽  
Hydrolysis Of

In the malaria parasite Plasmodium falciparum, erythrocytic trophozoites hydrolyse haemoglobin to provide amino acids for parasite protein synthesis. Cysteine protease inhibitors block parasite haemoglobin hydrolysis and development, indicating that cysteine proteases are required for these processes. Three papain-family cysteine protease sequences have been identified in the P. falciparum genome, but the specific roles of their gene products and other plasmodial proteases in haemoglobin hydrolysis are uncertain. Falcipain-2 was recently identified as a principal trophozoite cysteine protease and potential drug target. The present study characterizes the related P. falciparum cysteine protease falcipain-3. As is the case with falcipain-2, falcipain-3 is expressed by trophozoites and appears to be located within the food vacuole, the site of haemoglobin hydrolysis. Both proteases require a reducing environment and acidic pH for optimal activity, and both prefer peptide substrates with leucine at the P2 position. The proteases differ, however, in that falcipain-3 undergoes efficient processing to an active form only at acidic pH, is more active and stable at acidic pH, and has much lower specific activity against typical papain-family peptide substrates, but has greater activity against native haemoglobin. Thus falcipain-3 is a second P. falciparum haemoglobinase that is particularly suited for the hydrolysis of native haemoglobin in the acidic food vacuole. The redundancy of cysteine proteases may offer optimized hydrolysis of both native haemoglobin and globin peptides. Consideration of both proteases will be necessary to evaluate cysteine protease inhibitors as antimalarial drugs.

scholarly journals Drug Repurposing for the SARS-CoV-2 Papain-Like Protease

2021 ◽  
Author(s):  
Chia-Chuan D Cho ◽  
Shuhua G Li ◽  
Kai S Yang ◽  
Tyler J Lalonde ◽  
Ge Yu ◽  
...  
Keyword(s):  
Protease Inhibitors ◽  
Cysteine Protease ◽  
Drug Repurposing ◽  
Cysteine Proteases ◽  
Host Cells ◽  
Nonstructural Proteins ◽  
Allosteric Site ◽  
Cysteine Protease Inhibitors ◽  
Ic50 Value ◽  
Depth Analysis

As the pathogen of COVID-19, SARS-CoV-2 encodes two essential cysteine proteases that process the pathogen's two large polypeptide translates ORF1a and ORF1ab in human host cells to form 15 functionally important, mature nonstructural proteins. One of the two enzymes, papain-like protease or PLpro, also possesses deubiquitination and deISGylation activities that suppresses host innate immune responses toward SARS-CoV-2 infection. Therefore, PLpro is a potential COVID-19 drug target. To repurpose drugs for PLpro, we experimentally screened 33 deubiquitinase and 37 cysteine protease inhibitors on their inhibition of PLpro. Our results showed that 15 deubiquitinase and 1 cysteine protease inhibitors exhibit potent inhibition of PLpro at 200 uM. More comprehensive characterizations revealed 7 inhibitors GRL0617, SJB2-043, TCID, DUB-IN-1, DUB-IN-3, PR-619, and S130 with an IC50 value below 60 uM and four inhibitors GRL0617, SJB2-043, TCID, and PR-619 with an IC50 value below 10 uM. Among four inhibitors with an IC50 value below 10 uM, SJB2-043 is the most unique in that it doesn't fully inhibit PLpro but has an outstanding IC50 value of 0.56 uM. SJB2-043 likely binds to an allosteric site of PLpro to convene its inhibition effect, which needs to be further investigated. As a pilot study, the current work indicates that COVID-19 drug repurposing by targeting PLpro holds promises but in-depth analysis of repurposed drugs is necessary to avoid omitting allosteric inhibitors.

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