Journal articles on the topic 'Cysteine protease'
To see the other types of publications on this topic, follow the link: Cysteine protease.
Create a spot-on reference in APA, MLA, Chicago, Harvard, and other styles
Consult the top 50 journal articles for your research on the topic 'Cysteine protease.'
Next to every source in the list of references, there is an 'Add to bibliography' button. Press on it, and we will generate automatically the bibliographic reference to the chosen work in the citation style you need: APA, MLA, Harvard, Chicago, Vancouver, etc.
You can also download the full text of the academic publication as pdf and read online its abstract whenever available in the metadata.
Browse journal articles on a wide variety of disciplines and organise your bibliography correctly.
1
Mitchell, Angela M., and R. Jude Samulski. "Mechanistic Insights into the Enhancement of Adeno-Associated Virus Transduction by Proteasome Inhibitors." Journal of Virology 87, no. 23 (September 11, 2013): 13035–41. http://dx.doi.org/10.1128/jvi.01826-13.
Full textAbstract:
Proteasome inhibitors (e.g., bortezomib, MG132) are known to enhance adeno-associated virus (AAV) transduction; however, whether this results from pleotropic proteasome inhibition or off-target serine and/or cysteine protease inhibition remains unresolved. Here, we examined recombinant AAV (rAAV) effects of a new proteasome inhibitor, carfilzomib, which specifically inhibits chymotrypsin-like proteasome activity and no other proteases. We determined that proteasome inhibitors act on rAAV through proteasome inhibition and not serine or cysteine protease inhibition, likely through positive changes late in transduction.
2
FAMPA, P., C. V. LISBOA, A. M. JANSEN, A. L. S. SANTOS, and M. I. RAMIREZ. "Protease expression analysis in recently field-isolated strains of Trypanosoma cruzi: a heterogeneous profile of cysteine protease activities between TC I and TC II major phylogenetic groups." Parasitology 135, no. 9 (July 14, 2008): 1093–100. http://dx.doi.org/10.1017/s0031182008004587.
Full textAbstract:
SUMMARYProtease expression among TCI and TCII field isolates was analysed. Gelatin-containing gels revealed hydrolysis bands with molecular masses ranging from 45 to 66 kDa. The general protease expression profile showed that TCII isolates presented higher heterogeneity compared to TCI. By utilizing protease inhibitors, we showed that all active proteases at acid pH are cysteine-proteases and all proteases active at alkaline pH are metalloproteases. However, the expression of cruzipain, the T. cruzi major cysteine-protease, did not reproduce a heterogeneous TCII cysteine zymogram profile. Dendogram analyses based on presence/absence matrices of proteases and cruzipain bands showed a TCI separation from the TCII group with 50–60% similarity. We suggest that the observed cysteine protease diversification contributes to differential host infection between TCI and II genotypes.
3
Jairajpuri, Mohamad Aman, and Shoyab Ansari. "Using serpins cysteine protease cross-specificity to possibly trap SARS-CoV-2 Mpro with reactive center loop chimera." Clinical Science 134, no. 17 (September 1, 2020): 2235–41. http://dx.doi.org/10.1042/cs20200767.
Full textAbstract:
Abstract Human serine protease inhibitors (serpins) are the main inhibitors of serine proteases, but some of them also have the capability to effectively inhibit cysteine proteases. Severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) main protease (Mpro) is a chymotrypsin-type cysteine protease that is needed to produce functional proteins essential for virus replication and transcription. Serpin traps its target proteases by presenting a reactive center loop (RCL) as protease-specific cleavage site, resulting in protease inactivation. Mpro target sites with its active site serine and other flanking residues can possibly interact with serpins. Alternatively, RCL cleavage site of serpins with known evidence of inhibition of cysteine proteases can be replaced by Mpro target site to make chimeric proteins. Purified chimeric serpin can possibly inhibit Mpro that can be assessed indirectly by observing the decrease in ability of Mpro to cleave its chromogenic substrate. Chimeric serpins with best interaction and active site binding and with ability to form 1:1 serpin–Mpro complex in human plasma can be assessed by using SDS/PAGE and Western blot analysis with serpin antibody. Trapping SARS-CoV-2 Mpro cysteine protease using cross-class serpin cysteine protease inhibition activity is a novel idea with significant therapeutic potential.
4
SIJWALI, Puran S., Bhaskar R. SHENAI, Jiri GUT, Ajay SINGH, and Philip J. ROSENTHAL. "Expression and characterization of the Plasmodium falciparum haemoglobinase falcipain-3." Biochemical Journal 360, no. 2 (November 26, 2001): 481–89. http://dx.doi.org/10.1042/bj3600481.
Full textAbstract:
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.
5
Van Wyk, S. G., K. J. Kunert, B. J. Vorster, and U. Schluter. "Interaction of cysteine protease inhibitor mutants with cysteine proteases." South African Journal of Botany 76, no. 2 (April 2010): 406. http://dx.doi.org/10.1016/j.sajb.2010.02.053.
Full text
6
Semenov, Andrey, Jed E. Olson, and Philip J. Rosenthal. "Antimalarial Synergy of Cysteine and Aspartic Protease Inhibitors." Antimicrobial Agents and Chemotherapy 42, no. 9 (September 1, 1998): 2254–58. http://dx.doi.org/10.1128/aac.42.9.2254.
Full textAbstract:
ABSTRACT It has been proposed that the Plasmodium falciparumcysteine protease falcipain and aspartic proteases plasmepsin I and plasmepsin II act cooperatively to hydrolyze hemoglobin as a source of amino acids for erythrocytic parasites. Inhibitors of each of these proteases have potent antimalarial effects. We have now evaluated the antimalarial effects of combinations of cysteine and aspartic protease inhibitors. When incubated with cultured P. falciparumparasites, cysteine and aspartic protease inhibitors exhibited synergistic effects in blocking parasite metabolism and development. The inhibitors also demonstrated apparent synergistic inhibition of plasmodial hemoglobin degradation both in culture and in a murine malaria model. When evaluated for the treatment of murine malaria, a combination of cysteine and aspartic protease inhibitors was much more effective than higher concentrations of either compound used alone. These results support a model whereby plasmodial cysteine and aspartic proteases participate in the degradation of hemoglobin, and they suggest that combination antimalarial therapy with inhibitors of the two classes of proteases is worthy of further study.
7
Mann, Krin, and Hélène Sanfaçon. "Expanding Repertoire of Plant Positive-Strand RNA Virus Proteases." Viruses 11, no. 1 (January 15, 2019): 66. http://dx.doi.org/10.3390/v11010066.
Full textAbstract:
Many plant viruses express their proteins through a polyprotein strategy, requiring the acquisition of protease domains to regulate the release of functional mature proteins and/or intermediate polyproteins. Positive-strand RNA viruses constitute the vast majority of plant viruses and they are diverse in their genomic organization and protein expression strategies. Until recently, proteases encoded by positive-strand RNA viruses were described as belonging to two categories: (1) chymotrypsin-like cysteine and serine proteases and (2) papain-like cysteine protease. However, the functional characterization of plant virus cysteine and serine proteases has highlighted their diversity in terms of biological activities, cleavage site specificities, regulatory mechanisms, and three-dimensional structures. The recent discovery of a plant picorna-like virus glutamic protease with possible structural similarities with fungal and bacterial glutamic proteases also revealed new unexpected sources of protease domains. We discuss the variety of plant positive-strand RNA virus protease domains. We also highlight possible evolution scenarios of these viral proteases, including evidence for the exchange of protease domains amongst unrelated viruses.
8
Shibao, Priscila Yumi Tanaka, Milene Ferro, Fernando Fonseca Pereira de Paula, Bruno Salata Lima, and Flávio Henrique-Silva. "Identification and Functional Analysis of a Pseudo-Cysteine Protease from the Midgut Transcriptome of Sphenophorus levis." International Journal of Molecular Sciences 22, no. 21 (October 25, 2021): 11476. http://dx.doi.org/10.3390/ijms222111476.
Full textAbstract:
The Sphenophorus levis (Coleoptera, Curculionidae) is one of the main pests of sugarcane in Brazil. Although its major digestive proteases are known, its complex digestive process still needs to be further understood. We constructed a transcriptome from the midgut of 30-day-old larvae and identified sequences similar to its major digestive protease (cysteine cathepsin Sl-CathL), however, they presented a different amino acid than cysteine in the active cleft. We identified, recombinantly produced, and characterized Sl-CathL-CS, a pseudo cysteine protease, and verified that higher gene expression levels of Sl-CathL-CS occur in the midgut of 30-day old larvae. We reverted the serine residue to cysteine and compared the activity of the mutant (Sl-CathL-mutSC) with Sl-CathL-CS. Sl-CathL-CS presented no protease activity, but Sl-CathL-mutSC hydrolyzed Z-Phe-Arg-AMC (Vmax = 1017.60 ± 135.55, Km = 10.77 mM) and was inhibited by a cysteine protease inhibitor E-64 (Ki = 38.52 ± 1.20 μM), but not by the serine protease inhibitor PMSF. Additionally, Sl-CathL-CS interacted with a sugarcane cystatin, while Sl-CathL-mutSC presented weaker interaction. Finally, protein ligand docking reinforced the differences in the catalytic sites of native and mutant proteins. These results indicate that Sl-CathL-CS is a pseudo-cysteine protease that assists protein digestion possibly by interacting with canecystatins, allowing the true proteases to work.
9
Lee, Jung-Yub, Su-Min Song, Eun-Kyung Moon, Yu-Ran Lee, Bijay Kumar Jha, Dinzouna-Boutamba Sylvatrie Danne, Hee-Jae Cha, et al. "Cysteine Protease Inhibitor (AcStefin) Is Required for Complete Cyst Formation of Acanthamoeba." Eukaryotic Cell 12, no. 4 (February 8, 2013): 567–74. http://dx.doi.org/10.1128/ec.00308-12.
Full textAbstract:
ABSTRACTThe encystation ofAcanthamoebaleads to the formation of resilient cysts from vegetative trophozoites. This process is essential for parasite survival under unfavorable conditions, such as those associated with starvation, low temperatures, and biocides. Furthermore, cysteine proteases have been implicated in the massive turnover of intracellular components required for encystation. Thus, strict modulation of the activities of cysteine proteases is required to protectAcanthamoebafrom intracellular damage. However, mechanisms underlying the control of protease activity during encystation have not been established inAcanthamoeba. In the present study, we identified and characterizedAcanthamoebacysteine protease inhibitor (AcStefin), which was found to be highly expressed during encystation and to be associated with lysosomes by fluorescence microscopy. Recombinant AcStefin inhibited various cysteine proteases, including human cathepsin B, human cathepsin L, and papain. Transfection with small interfering RNA against AcStefin increased cysteine protease activity during encystation and resulted in incomplete cyst formation, reduced excystation efficiency, and a significant reduction in cytoplasmic area. Taken together, these results indicate that AcStefin is involved in the modulation of cysteine proteases and that it plays an essential role during the encystation ofAcanthamoeba.
10
Davis, David A., Keisuke Yusa, Laura A. Gillim, Fonda M. Newcomb, Hiroaki Mitsuya, and Robert Yarchoan. "Conserved Cysteines of the Human Immunodeficiency Virus Type 1 Protease Are Involved in Regulation of Polyprotein Processing and Viral Maturation of Immature Virions." Journal of Virology 73, no. 2 (February 1, 1999): 1156–64. http://dx.doi.org/10.1128/jvi.73.2.1156-1164.1999.
Full textAbstract:
ABSTRACT We investigated the role of the two highly conserved cysteine residues, cysteines 67 and 95, of the human immunodeficiency virus type 1 (HIV-1) protease in regulating the activity of that protease during viral maturation. To this end, we generated four HIV-1 molecular clones: the wild type, containing both cysteine residues; a protease mutant in which the cysteine at position 67 was replaced by an alanine (C67A); a C95A protease mutant; and a double mutant (C67A C95A). When immature virions were produced in the presence of an HIV-1 protease inhibitor, KNI-272, and the inhibitor was later removed, limited polyprotein processing was observed for wild-type virion preparations over a 20-h period. Treatment of immature wild-type virions with the reducing agent dithiothreitol considerably improved the rate and extent of Gag processing, suggesting that the protease is, in part, reversibly inactivated by oxidation of the cysteine residues. In support of this, C67A C95A virions processed Gag up to fivefold faster than wild-type virions in the absence of a reducing agent. Furthermore, oxidizing agents, such as H2O2 and diamide, inhibited Gag processing of wild-type virions, and this effect was dependent on the presence of cysteine 95. Electron microscopy revealed that a greater percentage of double-mutant virions than wild-type virions developed a mature-like morphology on removal of the inhibitor. These studies provide evidence that under normal culture conditions the cysteines of the HIV-1 protease are susceptible to oxidation during viral maturation, thus preventing immature virions from undergoing complete processing following their release. This is consistent with the cysteines being involved in the regulation of viral maturation in cells under oxidative stress.
11
López, D., and M. Del Val. "Selective involvement of proteasomes and cysteine proteases in MHC class I antigen presentation." Journal of Immunology 159, no. 12 (December 15, 1997): 5769–72. http://dx.doi.org/10.4049/jimmunol.159.12.5769.
Full textAbstract:
Abstract CTL recognize peptides derived from protein Ags bound to MHC-class I molecules. Proteasomes probably participate in the generation of these peptide epitopes. We investigated the role of proteasomes in the presentation of endogenously synthesized short viral proteins. To this end, we employed proteasome and cysteine protease inhibitors and two closely related recombinant vaccinia viruses that code for 17- and 19-amino acid-long products encompassing murine CMV 9pp89 epitope. Presentation of both minigene products required processing to shorter peptides and was independent of ubiquitination. Proteasomes were necessary for processing the 17-mer product, and cysteine proteases were not required. In contrast, the 19-mer product could be processed in parallel either by proteasomes or by cysteine proteases independently. These results highlight the diversity of alternative processing pathways even for short peptidic Ags, provide evidence for the involvement of cysteine proteases in MHC class I presentation, and show that cleavage by cysteine proteases is governed by sequences flanking the epitope.
12
Her, Kyu-Hee, Kwang-Sig KIM, Gukmyung Choi, Seung Hyoung Kim, and Young-Bae Chung. "Partial characterization of a cysteine protease from Panonychus citri." Journal of Medicine and Life Science 4, no. 1 (June 1, 2006): 73–75. http://dx.doi.org/10.22730/jmls.2006.4.1.73.
Full textAbstract:
A cysteine protease activity of Panonychus citri was detected and partially characterized. Proteolytic activity of the enzyme was enhanced by the addition of reducing agent, dithiothreitol (DTT), and also it was absolutely inhibited by cysteine protease specific inhibitors, such as trans-epoxy-succinyl-L-leucyl-amido (4-guanidino) butane (E.64) and iodoacetic acid (IAA). The other specific inhibitors of serine- and metallo-proteases could not inhibit the enzyme activity. This result showed for the first time cysteine protease activity of P. citri. Further studies on the biological roles of the enzyme are required.
13
Cusack, M., A. G. Stephen, R. Powls, and R. J. Beynon. "Purification and characterization of thaumatopain, a cysteine protease from the arils of the plant Thaumatococcus daniellii." Biochemical Journal 274, no. 1 (February 15, 1991): 231–36. http://dx.doi.org/10.1042/bj2740231.
Full textAbstract:
Aqueous extracts of the aril of the seed of Thaumatococcus daniellii contain, in addition to the intensely sweet protein thaumatin, a cysteine protease that we have termed thaumatopain. Thaumatopain has been purified by ion-exchange chromatography from arils, and is a monomeric protein of Mr 30,000. The protease strongly resembles papain in proteolytic activity, pH optima, susceptibility to inhibitors of cysteine proteases and in N-terminal sequence. The protease has also been identified in crude aril extracts by affinity labelling with iodo[14C]acetate. Thaumatopain is responsible for the cysteine protease activity previously attributed to thaumatin. Thaumatin is digested by thaumatopain at neutral to alkaline pH values.
14
Parikh, Sunil, Jun Liu, Puran Sijwali, Jiri Gut, Daniel E. Goldberg, and Philip J. Rosenthal. "Antimalarial Effects of Human Immunodeficiency Virus Type 1 Protease Inhibitors Differ from Those of the Aspartic Protease Inhibitor Pepstatin." Antimicrobial Agents and Chemotherapy 50, no. 6 (June 2006): 2207–9. http://dx.doi.org/10.1128/aac.00022-06.
Full textAbstract:
ABSTRACT Human immunodeficiency virus type 1 protease inhibitors (HIVPIs) and pepstatin are aspartic protease inhibitors with antimalarial activity. In contrast to pepstatin, HIVPIs were not synergistic with a cysteine protease inhibitor or more active against parasites with the cysteine protease falcipain-2 knocked out than against wild-type parasites. As with pepstatin, HIVPIs were equally active against wild-type parasites and against parasites with the food vacuole plasmepsin aspartic proteases knocked out. The antimalarial mechanism of HIVPIs differs from that of pepstatin.
15
Renko, Miha, Tanja Zupan, David F. Plaza, Stefanie S. Schmieder, Milica Perišić Nanut, Janko Kos, Dušan Turk, Markus Künzler, and Jerica Sabotič. "Cocaprins, β-trefoil Fold Inhibitors of Cysteine and Aspartic Proteases from Coprinopsis cinerea." International Journal of Molecular Sciences 23, no. 9 (April 28, 2022): 4916. http://dx.doi.org/10.3390/ijms23094916.
Full textAbstract:
We introduce a new family of fungal protease inhibitors with β-trefoil fold from the mushroom Coprinopsis cinerea, named cocaprins, which inhibit both cysteine and aspartic proteases. Two cocaprin-encoding genes are differentially expressed in fungal tissues. One is highly transcribed in vegetative mycelium and the other in the stipes of mature fruiting bodies. Cocaprins are small proteins (15 kDa) with acidic isoelectric points that form dimers. The three-dimensional structure of cocaprin 1 showed similarity to fungal β-trefoil lectins. Cocaprins inhibit plant C1 family cysteine proteases with Ki in the micromolar range, but do not inhibit the C13 family protease legumain, which distinguishes them from mycocypins. Cocaprins also inhibit the aspartic protease pepsin with Ki in the low micromolar range. Mutagenesis revealed that the β2-β3 loop is involved in the inhibition of cysteine proteases and that the inhibitory reactive sites for aspartic and cysteine proteases are located at different positions on the protein. Their biological function is thought to be the regulation of endogenous proteolytic activities or in defense against fungal antagonists. Cocaprins are the first characterized aspartic protease inhibitors with β-trefoil fold from fungi, and demonstrate the incredible plasticity of loop functionalization in fungal proteins with β-trefoil fold.
16
Ge, Zhao-Yu, Pin-Jun Wan, Guo-Qing Li, Yong-gui Xia, and Zhao-Jun Han. "Characterization of cysteine protease-like genes in the striped rice stem borer, Chilo suppressalis." Genome 57, no. 2 (February 2014): 79–88. http://dx.doi.org/10.1139/gen-2013-0188.
Full textAbstract:
The striped rice stem borer, Chilo suppressalis (Walker), is a major pest for rice production in China and the rest of Southeast Asia. Chemical control is the main means to alleviate losses due to this pest, which causes serious environmental pollution. An effective and environmentally friendly approach is needed for the management of the striped rice stem borer. Cysteine proteases in insects could be useful targets for pest management either through engineering plant protease inhibitors, targeting insect digestive cysteine proteases, or through RNA interference-based silencing of cysteine proteases, disrupting developmental regulation of insects. In this study, eight cysteine protease-like genes were identified and partially characterized. The genes CCO2 and CCL4 were exclusively expressed in the larval gut, and their expression was affected by the state of nutrition in the insect. The expression of CCL2, CCL3, and CCO1 was significantly affected by the type of host plant, suggesting a role in host plant – insect interactions. Our initial characterization of the striped rice stem borer cysteine protease-like genes provides a foundation for further research on this important group of genes in this major insect pest of rice.
17
Dubin, G., J. Stec-Niemczyk, T. Dylag, J. Silberring, A. Dubin, and J. Potempa. "Characterisation of a highly specific, endogenous inhibitor of cysteine protease from Staphylococcus epidermidis, a new member of the staphostatin family." Biological Chemistry 385, no. 6 (June 7, 2004): 543–46. http://dx.doi.org/10.1515/bc.2004.064.
Full textAbstract:
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.
18
Brooks, D. R., L. Tetley, G. H. Coombs, and J. C. Mottram. "Processing and trafficking of cysteine proteases in Leishmania mexicana." Journal of Cell Science 113, no. 22 (November 15, 2000): 4035–41. http://dx.doi.org/10.1242/jcs.113.22.4035.
Full textAbstract:
Removal of the pro-domain of a cysteine protease is essential for activation of the enzyme. We have engineered a cysteine protease (CPB2.8) of the protozoan parasite Leishmania mexicana by site-directed mutagenesis to remove the active site cysteine (to produce CPB(C25G)). When CPB(C25G) was expressed in a L. mexicana mutant lacking all CPB genes, the inactive pro-enzyme was processed to the mature protein and trafficked to the lysosome. These results show that auto-activation is not required for correct processing of CPB in vivo. When CPB(C25G) was expressed in a L. mexicana mutant lacking both CPA and CPB genes, the majority of the pro-enzyme remained unprocessed and accumulated in the flagellar pocket. These data reveal that CPA can directly or indirectly process CPB(C25G) and suggest that cysteine proteases are targeted to lysosomes via the flagellar pocket. Moreover, they show that another protease can process CPB in the absence of either CPA or CPB, albeit less efficiently. Abolition of the glycosylation site in the mature domain of CPB did not affect enzyme processing, targeting or in vitro activity towards gelatin. This indicates that glycosylation is not required for trafficking. Together these findings provide evidence that the major route of trafficking of Leishmania cysteine proteases to lysosomes is via the flagellar pocket and therefore differs significantly from cysteine protease trafficking in mammalian cells.
19
Aly, Ahmed S. I., and Kai Matuschewski. "A malarial cysteine protease is necessary for Plasmodium sporozoite egress from oocysts." Journal of Experimental Medicine 202, no. 2 (July 18, 2005): 225–30. http://dx.doi.org/10.1084/jem.20050545.
Full textAbstract:
The Plasmodium life cycle is a sequence of alternating invasive and replicative stages within the vertebrate and invertebrate hosts. How malarial parasites exit their host cells after completion of reproduction remains largely unsolved. Inhibitor studies indicated a role of Plasmodium cysteine proteases in merozoite release from host erythrocytes. To validate a vital function of malarial cysteine proteases in active parasite egress, we searched for target genes that can be analyzed functionally by reverse genetics. Herein, we describe a complete arrest of Plasmodium sporozoite egress from Anopheles midgut oocysts by targeted disruption of a stage-specific cysteine protease. Our findings show that sporozoites exit oocysts by parasite-dependent proteolysis rather than by passive oocyst rupture resulting from parasite growth. We provide genetic proof that malarial cysteine proteases are necessary for egress of invasive stages from their intracellular compartment and propose that similar cysteine protease–dependent mechanisms occur during egress from liver-stage and blood-stage schizonts.
20
Wileman, T., L. P. Kane, and C. Terhorst. "Degradation of T-cell receptor chains in the endoplasmic reticulum is inhibited by inhibitors of cysteine proteases." Cell Regulation 2, no. 9 (September 1991): 753–65. http://dx.doi.org/10.1091/mbc.2.9.753.
Full textAbstract:
The endoplasmic reticulum, or an organelle closely associated with it, contains proteases that can be used to remove partially assembled or improperly folded proteins. Very little is known at present about the types of protease that degrade these proteins. The beta chain and cluster of differentiation (CD)3 delta subunit of the human T-cell antigen receptor (TCR) are degraded shortly after synthesis. In this study Chinese hamster ovary (CHO) cells transfected with either beta or delta were incubated with a panel of protease inhibitors, and the rates of degradation of the transfected proteins were followed using chain-specific enzyme-linked immunosorbent assays (ELISAs). Of the protease inhibitors tested, degradation of both chains was highly sensitive to sulfhydryl reagents and peptidyl inhibitors of cysteine proteases. Concentrations of inhibitors that produced near complete inhibition of degradation in the endoplasmic reticulum did not cause gross changes in cellular ATP levels nor did they significantly slow constitutive secretion from CHO cells. The inhibitors did not affect the ability of CHO cells to synthesize and assemble disulphide-linked TCR zeta dimers. We conclude that the protease inhibitors were not toxic to cells and did not affect the biosynthetic activity of the endoplasmic reticulum. Furthermore, they did not alter the ability of the endoplasmic reticulum to deliver its content to the Golgi apparatus. Taken together, these results suggest that the cysteine protease inhibitors slow degradation in the endoplasmic reticulum through an action on cysteine proteases. The results imply that the endoplasmic reticulum contains cysteine proteases that can be used to remove retained proteins.
21
McGlinchey, Ryan P., and Jennifer C. Lee. "Cysteine cathepsins are essential in lysosomal degradation of α-synuclein." Proceedings of the National Academy of Sciences 112, no. 30 (July 13, 2015): 9322–27. http://dx.doi.org/10.1073/pnas.1500937112.
Full textAbstract:
A cellular feature of Parkinson’s disease is cytosolic accumulation and amyloid formation of α-synuclein (α-syn), implicating a misregulation or impairment of protein degradation pathways involving the proteasome and lysosome. Within lysosomes, cathepsin D (CtsD), an aspartyl protease, is suggested to be the main protease for α-syn clearance; however, the protease alone only generates amyloidogenic C terminal-truncated species (e.g., 1–94, 5–94), implying that other proteases and/or environmental factors are needed to facilitate degradation and to avoid α-syn aggregation in vivo. Using liquid chromatography–mass spectrometry, to our knowledge, we report the first peptide cleavage map of the lysosomal degradation process of α-syn. Studies of purified mouse brain and liver lysosomal extracts and individual human cathepsins demonstrate a direct involvement of cysteine cathepsin B (CtsB) and L (CtsL). Both CtsB and CtsL cleave α-syn within its amyloid region and circumvent fibril formation. For CtsD, only in the presence of anionic phospholipids can this protease cleave throughout the α-syn sequence, suggesting that phospholipids are crucial for its activity. Taken together, an interplay exists between α-syn conformation and cathepsin activity with CtsL as the most efficient under the conditions examined. Notably, we discovered that CtsL efficiently degrades α-syn amyloid fibrils, which by definition are resistant to broad spectrum proteases. This work implicates CtsB and CtsL as essential in α-syn lysosomal degradation, establishing groundwork to explore mechanisms to enhance their cellular activity and levels as a potential strategy for clearance of α-syn.
22
Morton, P. A., M. L. Zacheis, K. S. Giacoletto, J. A. Manning, and B. D. Schwartz. "Delivery of nascent MHC class II-invariant chain complexes to lysosomal compartments and proteolysis of invariant chain by cysteine proteases precedes peptide binding in B-lymphoblastoid cells." Journal of Immunology 154, no. 1 (January 1, 1995): 137–50. http://dx.doi.org/10.4049/jimmunol.154.1.137.
Full textAbstract:
Abstract The intracellular trafficking, proteolysis, and dissociation of invariant chain (li) associated with nascent class II molecules was examined in B-lymphoblastoid cells. Metabolic labeling and Percoll gradient centrifugation was used to assess the kinetics of delivery and processing of class II-li complexes within the endocytic pathway. Catabolism of class II-li complexes rapidly followed their delivery from post-Golgi compartments to dense lysosome-like compartments distinct from early and late endosomes. Direct peptide binding assays revealed that class II molecules associated with even small N-terminal fragments of li failed to bind peptide. Cysteine protease inhibitors alone blocked li proteolysis/dissociation and accumulation of class II-li biosynthetic intermediates within lysosome-containing compartments. Active-site labeling of cysteine proteases in B cells was used to identify cysteine proteases capable of mediating li proteolysis within endosomal compartments. Our results indicate rapid, possibly direct, transport of nascent class II-li complexes from the Golgi/trans-Golgi network to dense lysosomal compartments wherein cysteine protease(s), likely including cathepsin B, mediate complete removal of li. Inhibition of cysteine protease activity results in the accumulation of incompletely processed class II-li complexes, which lack peptide binding ability, within lysosomal compartments.
23
Saitoh, Eiichi, Shinya Yamamoto, Eishiro Okamoto, Yoshimi Hayakawa, Takashi Hoshino, Ritsuko Sato, Satoko Isemura, Sadami Ohtsubo, and Masayuki Taniguchi. "Identification of Cysteine Proteases and Screening of Cysteine Protease Inhibitors in Biological Samples by a Two-Dimensional Gel System of Zymography and Reverse Zymography." Analytical Chemistry Insights 2 (January 2007): 117739010700200. http://dx.doi.org/10.4137/117739010700200011.
Full textAbstract:
We have developed a two-dimensional (2D-) gel system of zymography and reverse zymography for the detection and characterization of proteases and protease inhibitors. Isoelectric focusing (IEF) agarose gels with pH gradients were employed for separation in the first-dimension and sodium dodecyl sulfate (SDS)-polyacrylamide gel copolymerized with gelatin used for the second dimension. Proteases and protease inhibitors separated by IEF gel were applied on the second gel without trichloroacetic acid (TCA) fixation. Protease activity in the 2D-gel was visualized as transparent spots where gelatin substrate was digested after commassie brilliant blue (CBB) staining. Some of the transparent spots from the skin mucus extract of rainbow trout were determined to be a cysteine protease through use of E-64 or CA-074. In the reverse zymography technique, the gel was incubated with papain solution at 37°C for 18 h. Cysteine protease inhibitors from broad bean seeds were detected as clear blue spots after CBB staining. The amino (N-) terminal sequences of four papain inhibitor spots thus detected were demonstrated to be identical to that of favin β chain, a broad bean lectin. Taken together, our system can be considered to be an efficient technique for discovering and characterizing new proteases and protease inhibitors in biological samples. This is the first report describing a 2D-gel system of zymography and reverse zymography.
24
Karlsson, Anna, Patricia Saravia-Otten, Karin Tegmark, Eva Morfeldt, and Staffan Arvidson. "Decreased Amounts of Cell Wall-Associated Protein A and Fibronectin-Binding Proteins in Staphylococcus aureus sarA Mutants due to Up-Regulation of Extracellular Proteases." Infection and Immunity 69, no. 8 (August 1, 2001): 4742–48. http://dx.doi.org/10.1128/iai.69.8.4742-4748.2001.
Full textAbstract:
ABSTRACT Data have been presented indicating that Staphylococcus aureus cell surface protein can be degraded by extracellular proteases produced by the same bacterium. We have found that insarA mutant cells, which produce high amounts of four major extracellular proteases (staphylococcal serine protease [V8 protease] [SspA], cysteine protease [SspB], aureolysin [metalloprotease] [Aur], and staphopain [Scp]), the levels of cell-bound fibronectin-binding proteins (FnBPs) and protein A were very low compared to those of wild-type cells, in spite of unaltered or increased transcription of the corresponding genes. Cultivation ofsarA mutant cells in the presence of the global protease inhibitor α2-macroglobulin resulted in a 16-fold increase in cell-bound FnBPs, indicating that extracellular proteases were responsible for the decreased amounts of FnBPs in sarAmutant cells. The protease inhibitor E64 had no effect on the level of FnBPs, indicating that cysteine proteases were not involved. Inactivation of either ssp or aur in the prototype S. aureus strain 8325-4 resulted in a threefold increase in the amount of cell-bound FnBPs. Inactivation of the same protease genes in a sarA mutant of 8325-4 resulted in a 10- to 20-fold increase in cell-bound protein A. As the serine protease requires aureolysin to be activated, it can thus be concluded that the serine protease is the most important protease in the release of cell-bound FnBPs and protein A.
25
Hook, V. Y. H., and S. R. Hwang. "Novel Secretory Vesicle Serpins, Endopin 1 and Endopin 2: Endogenous Protease Inhibitors with Distinct Target Protease Specificities." Biological Chemistry 383, no. 7-8 (August 27, 2002): 1067–74. http://dx.doi.org/10.1515/bc.2002.115.
Full textAbstract:
Abstract Secretory vesicles of neuroendocrine cells possess multiple proteases for proteolytic processing of proteins into biologically active peptide components, such as peptide hormones and neurotransmitters. The importance of proteases within secretory vesicles predicts the presence of endogenous protease inhibitors in this subcellular compartment. Notably, serpins represent a diverse class of endogenous protease inhibitors that possess selective target protease specificities, defined by the reactive site loop domains (RSL). In the search for endogenous serpins in model secretory vesicles of neuroendocrine chromaffin cells, the presence of serpins related to α1-antichymotrypsin (ACT) was detected by Western blots with antiACT. Molecular cloning revealed the primary structures of two unique serpins, endopin 1 and endopin 2, that possess homology to ACT. Of particular interest was the observation that distinct RSL domains of these new serpins predicted that endopin 1 would inhibit trypsinlike serine proteases cleaving at basic residues, and endopin 2 would inhibit both elastase and papain that represent serine and cysteine proteases, respectively. Endopin 1 showed selective inhibition of trypsin, but did not inhibit chymotrypsin, elastase, or subtilisin. Endopin 2 demonstrated crossclass inhibition of the cysteine protease papain and the serine protease elastase. Endopin 2 did not inhibit chymotrypsin, trypsin, plasmin, thrombin, furin, or cathepsin B. Endopin 1 and endopin 2 each formed SDSstable complexes with target proteases, a characteristic property of serpins. In neuroendocrine chromaffin cells from adrenal medulla, endopin 1 and endopin 2 were both localized to secretory vesicles. Moreover, the inhibitory activity of endopin 2 was optimized under reducing conditions, which required reduced Cys-374; this property is consistent with the presence of endogenous reducing agents in secretory vesicles in vivo. These new findings demonstrate the presence of unique secretory vesicle serpins, endopin 1 and endopin 2, which possess distinct target protease selectivities. Endopin 1 inhibits trypsinlike proteases; endopin 2 possesses crossclass inhibition for inhibition of papainlike cysteine proteases and elastaselike serine proteases. It will be of interest in future studies to define the endogenous protease targets of these two novel secretory vesicle serpins.
26
PARAMÁ, A., R. IGLESIAS, M. F. ÁLVAREZ, J. LEIRO, F. M. UBEIRA, and M. L. SANMARTÍN. "Cysteine proteinase activities in the fish pathogenPhilasterides dicentrarchi(Ciliophora: Scuticociliatida)." Parasitology 128, no. 5 (May 2004): 541–48. http://dx.doi.org/10.1017/s0031182004004883.
Full textAbstract:
This study investigated protease activities in a crude extract andin vitroexcretion/secretion (E/S) products ofPhilasterides dicentrarchi, a ciliate fish parasite causing economically significant losses in aquaculture. Gelatin/SDS–PAGE analysis (pH 4, reducing conditions) detected 7 bands with gelatinolytic activity (approximate molecular weights 30–63 kDa) in the crude extract. The banding pattern observed in analysis of E/S products was practically identical, except for 1 low-molecular-weight band detected in the crude extract but not in the E/S products. In assays with synthetic peptidep-nitroanilide substrates, the crude extract hydrolysed substrates characteristic of cysteine proteases, namely Z-Arg-Arg pNA, Bz-Phe-Val-Arg pNA and Z-Phe-Arg pNA. These activities were strongly inhibited by the cysteine protease inhibitor E-64 and by Ac-Leu-Val-Lys aldehyde, a potent inhibitor of cysteine proteases of the cathepsin B protease subfamily. The proteases present in the crude extract degraded both type-I collagen and haemoglobinin vitro, consistent with roles in tissue invasion and nutrition respectively. Again, E-64 completely (collagen) or markedly (haemoglobin) inhibited this degradation. Finally, the histolytic activity of the ciliate in turbot fibroblast monolayers was strongly reduced in the presence of E-64, confirming the importance of secreted cysteine proteinases in the biology ofPhilasterides dicentrarchi.
27
Shompole, Sankale, and Douglas P. Jasmer. "Cathepsin B-like Cysteine Proteases Confer Intestinal Cysteine Protease Activity inHaemonchus contortus." Journal of Biological Chemistry 276, no. 4 (October 13, 2000): 2928–34. http://dx.doi.org/10.1074/jbc.m007321200.
Full text
28
Draper, Deborah, William Donohoe, Leo Mortimer, and R. Phillip Heine. "Cysteine Proteases ofTrichomonas vaginalisDegrade Secretory Leukocyte Protease Inhibitor." Journal of Infectious Diseases 178, no. 3 (September 1998): 815–19. http://dx.doi.org/10.1086/515366.
Full text
29
Sloan, Sarah, Caitlin Jenvey, Callum Cairns, and Michael Stear. "Cathepsin F of Teladorsagia circumcincta is a recently evolved cysteine protease." Evolutionary Bioinformatics 16 (January 2020): 117693432096252. http://dx.doi.org/10.1177/1176934320962521.
Full textAbstract:
Parasitic cysteine proteases are involved in parasite stage transition, invasion of host tissues, nutrient uptake, and immune evasion. The cysteine protease cathepsin F is the most abundant protein produced by fourth-stage larvae (L4) of the nematode Teladorsagia circumcincta, while its transcript is only detectable in L4 and adults. T. circumcincta cathepsin F is a recently evolved cysteine protease that does not fall clearly into either of the cathepsin L or F subfamilies. This protein exhibits characteristics of both cathepsins F and L, and its phylogenetic relationship to its closest homologs is distant, including proteins of closely related nematodes of the same subfamily.
30
Tušar, Livija, Aleksandra Usenik, Boris Turk, and Dušan Turk. "Mechanisms Applied by Protein Inhibitors to Inhibit Cysteine Proteases." International Journal of Molecular Sciences 22, no. 3 (January 20, 2021): 997. http://dx.doi.org/10.3390/ijms22030997.
Full textAbstract:
Protein inhibitors of proteases are an important tool of nature to regulate and control proteolysis in living organisms under physiological and pathological conditions. In this review, we analyzed the mechanisms of inhibition of cysteine proteases on the basis of structural information and compiled kinetic data. The gathered structural data indicate that the protein fold is not a major obstacle for the evolution of a protease inhibitor. It appears that nature can convert almost any starting fold into an inhibitor of a protease. In addition, there appears to be no general rule governing the inhibitory mechanism. The structural data make it clear that the “lock and key” mechanism is a historical concept with limited validity. However, the analysis suggests that the shape of the active site cleft of proteases imposes some restraints. When the S1 binding site is shaped as a pocket buried in the structure of protease, inhibitors can apply substrate-like binding mechanisms. In contrast, when the S1 binding site is in part exposed to solvent, the substrate-like inhibition cannot be employed. It appears that all proteases, with the exception of papain-like proteases, belong to the first group of proteases. Finally, we show a number of examples and provide hints on how to engineer protein inhibitors.
31
Zhu, Bingkuan, Fang Luo, Yi Shen, Wenbin Yang, Chengsong Sun, Jipeng Wang, Jian Li, et al. "Schistosoma japonicum cathepsin B2 (SjCB2) facilitates parasite invasion through the skin." PLOS Neglected Tropical Diseases 14, no. 10 (October 26, 2020): e0008810. http://dx.doi.org/10.1371/journal.pntd.0008810.
Full textAbstract:
Cercariae invasion of the human skin is the first step in schistosome infection. Proteases play key roles in this process. However, little is known about the related hydrolytic enzymes in Schistosoma japonicum. Here, we investigated the biochemical features, tissue distribution and biological roles of a cathepsin B cysteine protease, SjCB2, in the invasion process of S. japonicum cercariae. Enzyme activity analysis revealed that recombinant SjCB2 is a typical cysteine protease with optimum temperature and pH for activity at 37°C and 4.0, respectively, and can be totally inhibited by the cysteine protease inhibitor E-64. Immunoblotting showed that both the zymogen (50 kDa) and mature enzyme (30.5 kDa) forms of SjCB2 are expressed in the cercariae. It was observed that SjCB2 localized predominantly in the acetabular glands and their ducts of cercariae, suggesting that the protease could be released during the invasion process. The protease degraded collagen, elastin, keratin, fibronectin, immunoglobulin (A, G and M) and complement C3, protein components of the dermis and immune system. In addition, proteomic analysis demonstrated that SjCB2 can degrade the human epidermis. Furthermore, it was showed that anti-rSjCB2 IgG significantly reduced (22.94%) the ability of the cercariae to invade the skin. The cysteine protease, SjCB2, located in the acetabular glands and their ducts of S. japonicum cercariae. We propose that SjCB2 facilitates skin invasion by degrading the major proteins of the epidermis and dermis. However, this cysteine protease may play additional roles in host-parasite interaction by degrading immunoglobins and complement protein.
32
Lonsdale-Eccles, J. D., G. W. N. Mpimbaza, Z. R. M. Nkhungulu, J. Olobo, L. Smith, O. M. Tosomba, and D. J. Grab. "Trypanosomatid cysteine protease activity may be enhanced by a kininogen-like moiety from host serum." Biochemical Journal 305, no. 2 (January 15, 1995): 549–56. http://dx.doi.org/10.1042/bj3050549.
Full textAbstract:
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.
33
Monteiro, Ana C. S., Magnus Abrahamson, Ana P. C. A. Lima, Marcos A. Vannier-Santos, and Julio Scharfstein. "Identification, characterization and localization of chagasin, a tight-binding cysteine protease inhibitor inTrypanosoma cruzi." Journal of Cell Science 114, no. 21 (November 1, 2001): 3933–42. http://dx.doi.org/10.1242/jcs.114.21.3933.
Full textAbstract:
Lysosomal cysteine proteases from mammalian cells and plants are regulated by endogenous tight-binding inhibitors from the cystatin superfamily. The presence of cystatin-like inhibitors in lower eukaryotes such as protozoan parasites has not yet been demonstrated, although these cells express large quantities of cysteine proteases and may also count on endogenous inhibitors to regulate cellular proteolysis. Trypanosoma cruzi, the causative agent of Chagas’ heart disease, is a relevant model to explore this possibility because these intracellular parasites rely on their major lysosomal cysteine protease (cruzipain) to invade and multiply in mammalian host cells. Here we report the isolation, biochemical characterization, developmental stage distribution and subcellular localization of chagasin, an endogenous cysteine protease inhibitor in T. cruzi. We used high temperature induced denaturation to isolate a heat-stable cruzipain-binding protein (apparent molecular mass, 12 kDa) from epimastigote lysates. This protein was subsequently characterized as a tight-binding and reversible inhibitor of papain-like cysteine proteases. Immunoblotting indicated that the expression of chagasin is developmentally regulated and inversely correlated with that of cruzipain. Gold-labeled antibodies localized chagasin to the flagellar pocket and cytoplasmic vesicles of trypomastigotes and to the cell surface of amastigotes. Binding assays performed by probing living parasites with fluorescein (FITC)-cruzipain or FITC-chagasin revealed the presence of both inhibitor and protease at the cell surface of amastigotes. The intersection of chagasin and cruzipain trafficking pathways may represent a checkpoint for downstream regulation of proteolysis in trypanosomatid protozoa.
34
Serrano-Luna, José de Jesús, Isaac Cervantes-Sandoval, Jesús Calderón, Fernando Navarro-García, Victor Tsutsumi, and Mineko Shibayama. "Protease activities of Acanthamoeba polyphaga and Acanthamoeba castellanii." Canadian Journal of Microbiology 52, no. 1 (January 1, 2006): 16–23. http://dx.doi.org/10.1139/w05-114.
Full textAbstract:
Acanthamoeba spp. are free-living amoebae that cause amoebic granulomatous encephalitis, skin lesions, and ocular amoebic keratitis in humans. Several authors have suggested that proteases could play a role in the pathogenesis of these diseases. In the present work, we performed a partial biochemical characterization of proteases in crude extracts of Acanthamoeba spp. and in conditioned medium using 7.5% SDS–PAGE copolymerized with 0.1% m/v gelatin as substrate. We distinguished a total of 17 bands with proteolytic activity distributed in two species of Acanthamoeba. The bands ranged from 30 to 188 kDa in A. castellanii and from 34 to 144 kDa in A. polyphaga. Additionally, we showed that the pattern of protease activity differed in the two species of Acanthamoeba when pH was altered. By using protease inhibitors, we found that the proteolytic activities belonged mostly to the serine protease family and secondly to cysteine proteases and that the proteolytic activities from A. castellanii were higher than those in A. polyphaga. Furthermore, aprotinin was found to in hibit crude extract protease activity on Madin–Darby canine kidney (MDCK) monolayers. These data suggest that protease patterns could be more complex than previously reported.Key words: Acanthamoeba spp., amoebic keratitis, serine proteases, cysteine proteases, cytopathic effect.
35
Harding, C. V., J. France, R. Song, J. M. Farah, S. Chatterjee, M. Iqbal, and R. Siman. "Novel dipeptide aldehydes are proteasome inhibitors and block the MHC-I antigen-processing pathway." Journal of Immunology 155, no. 4 (August 15, 1995): 1767–75. http://dx.doi.org/10.4049/jimmunol.155.4.1767.
Full textAbstract:
Abstract Class I MHC (MHC-I) molecules present peptides derived from Ag that are processed in the cytosol. The proteasome is a multicatalytic protease complex that is present in the cytosol and has been implicated in cytosolic Ag processing. Novel dipeptide aldehydes were designed, synthesized, and demonstrated to specifically inhibit the chymotrypsin-like protease activity of isolated proteasomes, but produced relatively little inhibition of cathepsin B, a vacuolar cysteine protease. The inhibitors were membrane permeable and inhibited intracellular cleavage of a membrane-permeable fluorogenic substrate of the chymotrypsin-like proteasome activity. When a model Ag, OVA, was introduced into the cytoplasm of M12.B6 murine B cells by electroporation, the proteasome inhibitors blocked its processing for subsequent presentation by MHC-I molecules. The inhibitors had little effect on class II MHC processing of exogenous Ag. The potencies of different inhibitors for blockade of MHC-I Ag processing correlated directly with their potencies for inhibition of the chymotrypsin-like proteasome activity. In contrast, conventional inhibitors of vacuolar cysteine proteases (e.g., leupeptin and benzyloxycarbonyl-Phe-Ala-CHN2) had little effect on MHC-I processing or the chymotryspin-like activity of isolated proteasomes. These results directly demonstrate that inhibition of proteasome activity blocks MHC-I Ag processing, confirming a role for proteasomes in this pathway. Moreover, they suggest that the chymotrypsin-like activity of the proteasome may be of major importance to the cytosolic processing of at least some Ag.
36
NOURRISSON, C., I. WAWRZYNIAK, A. CIAN, V. LIVRELLI, E. VISCOGLIOSI, F. DELBAC, and P. POIRIER. "OnBlastocystissecreted cysteine proteases: a legumain-activated cathepsin B increases paracellular permeability of intestinal Caco-2 cell monolayers." Parasitology 143, no. 13 (September 9, 2016): 1713–22. http://dx.doi.org/10.1017/s0031182016001396.
Full textAbstract:
SUMMARYBlastocystisspp. pathogenic potential remains unclear as these anaerobic parasitic protozoa are frequently isolated from stools of both symptomatic and asymptomatic subjects.In silicoanalysis of the whole genome sequence ofBlastocystissubtype 7 revealed the presence of numerous proteolytic enzymes including cysteine proteases predicted to be secreted. To assess the potential impact of proteases on intestinal cells and gut function, we focused our study on two cysteine proteases, a legumain and a cathepsin B, which were previously identified inBlastocystissubtype 7 culture supernatants. Both cysteine proteases were produced as active recombinant proteins. Activation of the recombinant legumain was shown to be autocatalytic and triggered by acidic pH, whereas proteolytic activity of the recombinant cathepsin B was only recorded after co-incubation with the legumain. We then measured the diffusion of 4-kDa FITC-labelled dextran across Caco-2 cell monolayers following exposition to eitherBlastocystisculture supernatants or each recombinant protease. BothBlastocystisculture supernatants and recombinant activated cathepsin B induced an increase of Caco-2 cell monolayer permeability, and this effect was significantly inhibited by E-64, a specific cysteine protease inhibitor. Our results suggest that cathepsin B might play a role in pathogenesis ofBlastocystisby increasing intestinal cell permeability.
37
Vorster, Barend, Urte Schlüter, Magdeleen du Plessis, Stefan van Wyk, Matome Makgopa, Ignatious Ncube, Marian Quain, Karl Kunert, and Christine Foyer. "The Cysteine Protease–Cysteine Protease Inhibitor System Explored in Soybean Nodule Development." Agronomy 3, no. 3 (August 20, 2013): 550–70. http://dx.doi.org/10.3390/agronomy3030550.
Full text
38
Bevec, T., V. Stoka, G. Pungercic, I. Dolenc, and V. Turk. "Major histocompatibility complex class II-associated p41 invariant chain fragment is a strong inhibitor of lysosomal cathepsin L." Journal of Experimental Medicine 183, no. 4 (April 1, 1996): 1331–38. http://dx.doi.org/10.1084/jem.183.4.1331.
Full textAbstract:
The invariant chain (Ii) is associated with major histocompatibility complex class II molecules during early stages of their intracellular transport. In an acidic endosomal/lysosomal compartment, it is proteolytically cleaved and removed from class II heterodimers. Participation of aspartic and cysteine proteases has been observed in in vitro degradation of Ii, but the specific enzymes responsible for its in vivo processing are as yet undefined. We have previously isolated a noncovalent complex of the lysosomal cysteine protease cathepsin L with a peptide fragment derived from the p41 form of Ii from human kidney. Here we show that this Ii fragment, which is identical to the alternatively spliced segment of p41, is a very potent competitive inhibitor of cathepsin L (equilibrium inhibition constant Ki = 1.7 X 10(-12) M). It inhibits two other cysteine proteases, cathepsin H and papain, but to much lesser extent. Cysteine proteases cathepsins B, C, and S, as well as representatives of serine, aspartic, and metalloproteases, are not inhibited at all. These findings suggest a novel role for p41 in the regulation of various proteolytic activities during antigen processing and presentation. The Ii inhibitory fragment shows no sequence homology with the known cysteine protease inhibitors, and may, therefore, represent a new class.
39
Abdulkadir, Abdullahi, Adamu Yusuf Kabiru, Hadiza Lami Muhammad, Bala Alkali Mohammed, and Tawakaltu Abdulrasheed-Adeleke. "Partial Purification of Phytocystatin from Moringa oleifera lam." AROC in Pharmaceutical and Biotechnology 2, no. 1 (February 4, 2022): 09–17. http://dx.doi.org/10.53858/arocpb02010917.
Full textAbstract:
Background: Phytocystatins are plants cysteine protease inhibitors (CPIs) that are known for their numerous uses in medicine and biotechnology. Methods: Different extraction media which include, Sodium Hydroxide, Hydrochloric acid, Sodium Chloride, Sodium phosphate buffer and distilled Water were used to evaluate flower, leave, root, latex, and stem bark of Moringa oleifera for inhibitory activity against cysteine protease (Papain enzyme). The phosphate buffer extract with higher protease inhibitory activity was concentrated by cold acetone precipitation and further subjected to partial purification using ammonium sulphate fractionation and Hydrophobic chromatography on Phenyl Sepharose column. The molecular weight of partially purified protein was estimated by sodium dodecyl sulphate (SDS) polyacrylamide gel electrophoresis. Results: The extracts of Sodium phosphate buffer from the seeds of Moringa oleifera showed higher CPI activity of 11.23 units/mg protein. Cold acetone precipitated extract gave high yield of 94.6% protein with 60% inhibition of protease. The partially purified protein showed a fifty percent inhibitory concentration (IC50) of 1.22±0.3 mg/ml protein against the enzyme with 63% inhibition. An estimated molecular weight of 13.5kDa was obtained from electrophoretic analysis of partially purified protein. Conclusion: Moringa oleifera seeds could be a good source of low molecular weight protein Inhibitor of Cysteine protease and might be useful in biotechnology of traditional medicinal plants, in transgenic crops to arrest the negative pathogenic over expressions of cysteine proteases
40
Rümenapf, Tillmann, Robert Stark, Manuela Heimann, and Heinz-Jürgen Thiel. "N-Terminal Protease of Pestiviruses: Identification of Putative Catalytic Residues by Site-Directed Mutagenesis." Journal of Virology 72, no. 3 (March 1, 1998): 2544–47. http://dx.doi.org/10.1128/jvi.72.3.2544-2547.1998.
Full textAbstract:
ABSTRACT Pestiviruses are the only members of the Flaviviridaethat encode a nonstructural protease at the N terminus of their polyproteins. This N-terminal protease (Npro) cleaves itself off of the nascent polyprotein autocatalytically and thereby generates the N terminus of the adjacent viral capsid protein C. In previous reports, sequence similarities between Npro and the catalytic residues of papain-like cysteine proteases were put forward. To test this hypothesis, substitutions of cysteine and histidine residues within Npro were carried out by site-directed mutagenesis. Translation of the mutagenized Npro-C proteins in cell-free lysates confirmed that only the predicted Cys69 was an essential amino acid for proteolysis, not His130. Further essential residues were identified with His49 and Glu22. While it remains speculative whether Glu22-His49-Cys69 actually build a catalytic triad, these results invalidate the assumption that Npro is a papain-like cysteine protease.
41
Mootz, Joe M., Cheryl L. Malone, Lindsey N. Shaw, and Alexander R. Horswill. "Staphopains Modulate Staphylococcus aureus Biofilm Integrity." Infection and Immunity 81, no. 9 (June 24, 2013): 3227–38. http://dx.doi.org/10.1128/iai.00377-13.
Full textAbstract:
ABSTRACTStaphylococcus aureusis a known cause of chronic biofilm infections that can reside on medical implants or host tissue. Recent studies have demonstrated an important role for proteinaceous material in the biofilm structure. TheS. aureusgenome encodes many secreted proteases, and there is growing evidence that these enzymes have self-cleavage properties that alter biofilm integrity. However, the specific contribution of each protease and mechanism of biofilm modulation is not clear. To address this issue, we utilized a sigma factor B (ΔsigB) mutant where protease activity results in a biofilm-negative phenotype, thereby creating a condition where the protease(s) responsible for the phenotype could be identified. Using a plasma-coated microtiter assay, biofilm formation was restored to the ΔsigBmutant through the addition of the cysteine protease inhibitor E-64 or by using Staphostatin inhibitors that specifically target the extracellular cysteine proteases SspB and ScpA (called Staphopains). Through construction of gene deletion mutants, we determined that ansspB scpAdouble mutant restored ΔsigBbiofilm formation, and this recovery could be replicated in plasma-coated flow cell biofilms. Staphopain levels were also found to be decreased under biofilm-forming conditions, possibly allowing biofilm establishment. The treatment ofS. aureusbiofilms with purified SspB or ScpA enzyme inhibited their formation, and ScpA was also able to disperse an established biofilm. The antibiofilm properties of ScpA were conserved acrossS. aureusstrain lineages. These findings suggest an underappreciated role of the SspB and ScpA cysteine proteases in modulatingS. aureusbiofilm architecture.
42
Sallai, Roberto Carlos, Bruno Ramos Salu, Rosemeire Aparecida Silva-Lucca, Flávio Lopes Alves, Thiago Henrique Napoleão, Patrícia Maria Guedes Paiva, Rodrigo da Silva Ferreira, Misako Uemura Sampaio, and Maria Luiza Vilela Oliva. "Biotechnological Potential of Araucaria angustifolia Pine Nuts Extract and the Cysteine Protease Inhibitor AaCI-2S." Plants 9, no. 12 (November 30, 2020): 1676. http://dx.doi.org/10.3390/plants9121676.
Full textAbstract:
Protease inhibitors are involved in the regulation of endogenous cysteine proteases during seed development and play a defensive role because of their ability to inhibit exogenous proteases such as those present in the digestive tracts of insects. Araucaria angustifolia seeds, which can be used in human and animal feed, were investigated for their potential for the development of agricultural biotechnology and in the field of human health. In the pine nuts extract, which blocked the activities of cysteine proteases, it was detected potent insecticidal activity against termites (Nasutitermes corniger) belonging to the most abundant termite genus in tropical regions. The cysteine inhibitor (AaCI-2S) was purified by ion-exchange, size exclusion, and reversed-phase chromatography. Its functional and structural stability was confirmed by spectroscopic and circular dichroism studies, and by detection of inhibitory activity at different temperatures and pH values. Besides having activity on cysteine proteases from C. maculatus digestive tract, AaCI-2S inhibited papain, bromelain, ficin, and cathepsin L and impaired cell proliferation in gastric and prostate cancer cell lines. These properties qualify A. angustifolia seeds as a protein source with value properties of natural insecticide and to contain a protease inhibitor with the potential to be a bioactive molecule on different cancer cells.
43
Lorenzo, Mayra E., Jae U. Jung, and Hidde L. Ploegh. "Kaposi's Sarcoma-Associated Herpesvirus K3 Utilizes the Ubiquitin-Proteasome System in Routing Class I Major Histocompatibility Complexes to Late Endocytic Compartments." Journal of Virology 76, no. 11 (June 1, 2002): 5522–31. http://dx.doi.org/10.1128/jvi.76.11.5522-5531.2002.
Full textAbstract:
ABSTRACT Human herpesvirus 8 (HHV8) downregulates major histocompatibility complex (MHC) class I complexes from the plasma membrane via two of its genes, K3 and K5. The N termini of K3 and K5 contain a plant homeodomain (PHD) predicted to be structurally similar to RING domains found in E3 ubiquitin ligases. In view of the importance of the ubiquitin-proteasome system in sorting within the endocytic pathway, we analyzed its role in downregulation of MHC class I complexes in cells expressing K3. Proteasome inhibitors as well as cysteine and aspartyl protease inhibitors stabilize MHC class I complexes in cells expressing K3. However, proteasome inhibitors differentially affect sorting of MHC class I complexes within the endocytic pathway and prevent their delivery to a dense endosomal compartment. In this compartment, the cytoplasmic tail of MHC class I complexes is cleaved by cysteine proteases. The complex is then cleaved within the plane of the membrane by an aspartyl protease, resulting in a soluble MHC class I fragment composed of the lumenal domain of the heavy chain, β2-microglobulin (β2m), and peptide. We conclude that K3 not only directs internalization, but also targets MHC class I complexes to a dense endocytic compartment on the way to lysosomes in a ubiquitin-proteasome-dependent manner.
44
Strachecka, A. J., M. M. Gryzińska, M. Krauze, and K. Grzywnowicz. "Profile of the body surface proteolytic systém in Apis mellifera quee." Czech Journal of Animal Science 56, No. 1 (January 20, 2011): 15–22. http://dx.doi.org/10.17221/150/2009-cjas.
Full textAbstract:
The proteolytic system on the body surface of the honey bee has been insufficiently researched. In this study the body surface proteolytic activity was examined in queens at various developmental stages (eggs, larvae, pupae and imagines) in different seasons (spring, summer, autumn, winter). Extracts of the body surface material with water and detergent were used for an in vitro analysis of the proteolytic activity and protease inhibitor level assaying, as well as for an electrophoretic separation of the extracts in polyacrylamide gels. The following methods were used: protein content testing by the Lowry method (modified by Schacterle-Pollack), protease activity testing by the Anson method and protease inhibitor activity testing by the Lee and Lin method. Our studies revealed a high protease activity in an acidic environment (pH = 2.4; the material rinsed with detergent), as well as in neutral (pH = 7) and alkaline (pH = 11.2) environments (the material rinsed with water in both cases). The highest protein concentration values were observed in the imagines from summer. The lowest activities of the proteases and protease inhibitors were determined in the eggs from summer. The highest activities of the acidic, neutral and alkaline proteases were observed in the pupae from spring. The highest number of protease activity bands in PAGE zymography was obtained for the neutral and alkaline activities in the queens for all the seasons. In the queens all the catalytic protease types were present: asparagine and cysteine proteases at pH = 2.4; cysteine proteases and metalloproteases at pH = 7 and serine proteases at pH = 11.2. These results were crucial for the analysis of immunity mechanisms on the body surface of the honey bee.
45
Pandey, Kailash C., and Rajnikant Dixit. "Structure-Function of Falcipains: Malarial Cysteine Proteases." Journal of Tropical Medicine 2012 (2012): 1–11. http://dx.doi.org/10.1155/2012/345195.
Full textAbstract:
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.
46
Carrion, Ricardo, Young-Tae Ro, and Jean L. Patterson. "Purification, Identification, and Biochemical Characterization of a Host-Encoded Cysteine Protease That Cleaves a Leishmaniavirus Gag-Pol Polyprotein." Journal of Virology 77, no. 19 (October 1, 2003): 10448–55. http://dx.doi.org/10.1128/jvi.77.19.10448-10455.2003.
Full textAbstract:
ABSTRACT Leishmania RNA virus (LRV) is a double-stranded RNA virus that infects some strains of the protozoan parasite Leishmania. As with other totiviruses, LRV presumably expresses its polymerase by a ribosomal frameshift, resulting in a capsid-polymerase fusion protein. We have demonstrated previously that an LRV capsid-polymerase polyprotein is specifically cleaved by a Leishmania-encoded cysteine protease. This study reports the purification of this protease through a strategy involving anion-exchange chromatography and affinity chromatography. By using a Sepharose-immobilized lectin, concanavalin A, we isolated a fraction enriched with LRV polyprotein-specific protease activity. Analysis of the active fraction by sodium dodecyl sulfate-polyacrylamide gel electrophoreses and silver staining revealed a 50-kDa protein that, upon characterization by high-pressure liquid chromatography electrospray tandem mass spectrometry (electrospray ionization/MS/MS), was identified as a cysteine protease of trypanosomes. A partial amino acid sequence derived from the MS/MS data was compared with a protein database using BLAST software, revealing homology with several cysteine proteases of Leishmania and other trypanosomes. The protease exhibited remarkable temperature stability, while inhibitor studies characterized the protease as a trypsin-like cysteine protease—a novel finding for Leishmania. To elucidate substrate preferences, a panel of deletion mutations and single-amino-acid mutations were engineered into a Gag-Pol fusion construct that was subsequently transcribed and translated in vitro and then used in cleavage assays. The data suggest that there are a number of cleavage sites located within a 153-amino-acid region spanning both the carboxy-terminal capsid region and the amino-terminal polymerase domain, with LRV capsid exhibiting the greatest susceptibility to proteolysis.
47
Luhr, Katarina M., Elin K. Nordström, Peter Löw, Hans-Gustaf Ljunggren, Albert Taraboulos, and Krister Kristensson. "Scrapie Protein Degradation by Cysteine Proteases in CD11c+ Dendritic Cells and GT1-1 Neuronal Cells." Journal of Virology 78, no. 9 (May 1, 2004): 4776–82. http://dx.doi.org/10.1128/jvi.78.9.4776-4782.2004.
Full textAbstract:
ABSTRACT Dendritic cells (DC) of the CD11c+ myeloid phenotype have been implicated in the spread of scrapie in the host. Previously, we have shown that CD11c+ DC can cause a rapid degradation of proteinase K-resistant prion proteins (PrPSc) in vitro, indicating a possible role of these cells in the clearance of PrPSc. To determine the mechanisms of PrPSc degradation, CD11c+ DC that had been exposed to PrPSc derived from a neuronal cell line (GT1-1) infected with scrapie (ScGT1-1) were treated with a battery of protease inhibitors. Following treatment with the cysteine protease inhibitors (2S,3S)-trans-epoxysuccinyl-l-leucylamido-3-methylbutane (E-64c), its ethyl ester (E-64d), and leupeptin, the degradation of PrPSc was inhibited, while inhibitors of serine and aspartic and metalloproteases (aprotinin, pepstatin, and phosphoramidon) had no effect. An endogenous degradation of PrPSc in ScGT1-1 cells was revealed by inhibiting the expression of cellular PrP (PrPC) by RNA interference, and this degradation could also be inhibited by the cysteine protease inhibitors. Our data show that PrPSc is proteolytically cleaved preferentially by cysteine proteases in both CD11c+ DC and ScGT1-1 cells and that the degradation of PrPSc by proteases is different from that of PrPC. Interference by protease inhibitors with DC-induced processing of PrPSc has the potential to modify prion spread, clearance, and immunization in a host.
48
Rana, Sindhuprava, and R. Sivaperumal. "Virtual screening and molecular docking of Anti-Antileishmanial for selected pharmacophore for visceral Leishmaniasis." Journal of Drug Delivery and Therapeutics 8, no. 6-s (December 15, 2018): 230–35. http://dx.doi.org/10.22270/jddt.v8i6-s.2120.
Full textAbstract:
Objective: DNA amplification of Cysteine protease of Leishmania donovani and study the interaction of cysteine protease inhibitors, antileishmanial compounds with cysteine protease receptor in various computational programs.
Materials and methods: Cysteine protease DNA of Leishmania donovani was amplified by PCR. The sequence of cysteine protease has been modeled and docked with suitable inhibitors by using various servers and computational tools. The model was designed, compared and validated by DOPE and Verify 3D scores. The model and the compound interaction were studied by LibDock and other programs.
Results: Cysteine protease DNA of Leishmania donovani was successfully amplified by PCR. The structural modeling was done to achieve effective enzyme inhibition, inhibitors block the binding sites of that protein. Homology modeling of cysteine protease has been done and docked with suitable inhibitors by using various servers and computational tools. The model was designed, compared and validated by DOPE and Verify 3D scores by using DSv3.5. Licochalcone-a alone showed 37 LibDock conformations with 6 different poses, were suitably docked at the site 1 with hydrogen bond formation. The study would help to design the novel drugs in respect of resistant one for the treatment of harmful visceral Leishmaniasis.
Conclusion: The molecular interaction of vinyl sulfones, hydrazide derivatives, antileishmanial drugs molecules and carbohydrazide derivatives have exhibited ideal molecular interaction with cathepsin B, a cysteine protease of L. donovani, amino acids such as Cys29, Hisl88 and Asn208 has been found to be active residues. Licochalcone-a and hydrazide derivative may become future antileishmanial compounds, which needs to be tested in in vitro and in vivo.
Keywords: Cysteine Protease, Vinyl Sulfone, Hydrazide, Antileishmanial Drugs, Licochalcone, Visceral Leishmaniasis.
49
Smith, Katherine M., Alban Gaultier, Helene Cousin, Dominique Alfandari, Judith M. White, and Douglas W. DeSimone. "The cysteine-rich domain regulates ADAM protease function in vivo." Journal of Cell Biology 159, no. 5 (December 2, 2002): 893–902. http://dx.doi.org/10.1083/jcb.200206023.
Full textAbstract:
ADAMs are membrane-anchored proteases that regulate cell behavior by proteolytically modifying the cell surface and ECM. Like other membrane-anchored proteases, ADAMs contain candidate “adhesive” domains downstream of their metalloprotease domains. The mechanism by which membrane-anchored cell surface proteases utilize these putative adhesive domains to regulate protease function in vivo is not well understood. We address this important question by analyzing the relative contributions of downstream extracellular domains (disintegrin, cysteine rich, and EGF-like repeat) of the ADAM13 metalloprotease during Xenopus laevis development. When expressed in embryos, ADAM13 induces hyperplasia of the cement gland, whereas ADAM10 does not. Using chimeric constructs, we find that the metalloprotease domain of ADAM10 can substitute for that of ADAM13, but that specificity for cement gland expansion requires a downstream extracellular domain of ADAM13. Analysis of finer resolution chimeras indicates an essential role for the cysteine-rich domain and a supporting role for the disintegrin domain. These and other results reveal that the cysteine-rich domain of ADAM13 cooperates intramolecularly with the ADAM13 metalloprotease domain to regulate its function in vivo. Our findings thus provide the first evidence that a downstream extracellular adhesive domain plays an active role in regulating ADAM protease function in vivo. These findings are likely relevant to other membrane-anchored cell surface proteases.
50
Hooper, Nigel M. "Proteases: a primer." Essays in Biochemistry 38 (October 1, 2002): 1–8. http://dx.doi.org/10.1042/bse0380001.
Full textAbstract:
A protease can be defined as an enzyme that hydrolyses peptide bonds. Proteases can be divided into endopeptidases, which cleave internal peptide bonds in substrates, and exopeptidases, which cleave the terminal peptide bonds. Exopeptidases can be further subdivided into aminopeptidases and carboxypeptidases. The Schechter and Berger nomenclature provides a model for describing the interactions between the peptide substrate and the active site of a protease. Proteases can also be classified as aspartic proteases, cysteine proteases, metalloproteases, serine proteases and threonine proteases, depending on the nature of the active site. Different inhibitors can be used experimentally to distinguish between these classes of protease. The MEROPs database groups proteases into families on the basis of similarities in sequence and structure. Protease activity can be regulated in vivo by endogenous inhibitors, by the activation of zymogens and by altering the rate of their synthesis and degradation.