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The Journal of Clinical Pediatric... 2015To suggest Papacarie(®) as a new deproteinizing agent in comparison with indigenously prepared 10% papain gel before and after acid etching that may enhance the quality...
OBJECTIVE
To suggest Papacarie(®) as a new deproteinizing agent in comparison with indigenously prepared 10% papain gel before and after acid etching that may enhance the quality of the bond between enamel surface and composite resin complex.
STUDY DESIGN
One hundred and twenty five extracted human premolars were utilized and divided into five groups: In the group 1, enamel surface was etched and primer was applied. In group 2, treatment with papacarie(®) for 60 seconds followed by etching and primer application. In group 3, etching followed by treatment with papacarie(®) for 60 seconds and primer application. In group 4, treatment with 10% papain gel for 60 seconds followed by etching and primer application. In group 5, etching followed by treatment with 10% papain gel for 60 seconds and primer application . After bonding the brackets, the mechanical testing was performed using a Universal testing machine. The failure mode was analyzed using an adhesive remnant index. The etching patterns before and after application of papacarie(®) and 10% papain gel was also evaluated using SEM. The values obtained for shear bond strength were submitted to analysis of variance and Tukey test (p < 0.05).
RESULTS
It was observed that group 2 and group 4 had the highest shear bond strength and was statistically significant from other groups (p=0.001). Regarding Adhesive remnant index no statistical difference was seen between the groups (p=0.538).
CONCLUSION
Papacarie(®) or 10% papain gel can be used to deproteinize the enamel surface before acid etching to enhance the bond strength of orthodontic brackets.
Topics: Acid Etching, Dental; Adhesiveness; Dental Bonding; Dental Enamel; Dental Enamel Proteins; Dental Stress Analysis; Gels; Humans; Materials Testing; Microscopy, Electron, Scanning; Orthodontic Brackets; Papain; Phosphoric Acids; Protein Denaturation; Resin Cements; Shear Strength; Stress, Mechanical; Surface Properties
PubMed: 26161607
DOI: 10.17796/1053-4628-39.4.348 -
Allergology International : Official... Jun 2014Epicutaneous sensitization to allergens is important in the pathogenesis of not only skin inflammation such as atopic dermatitis but also "atopic march" in allergic...
BACKGROUND
Epicutaneous sensitization to allergens is important in the pathogenesis of not only skin inflammation such as atopic dermatitis but also "atopic march" in allergic diseases such as asthma and food allergies. We here examined antibody production and skin barrier dysfunction in mice epicutaneously administered papain, a plant-derived occupational allergen belonging to the same family of cysteine proteases as mite major group 1 allergens.
METHODS
Papain and Staphylococcus aureus V8 protease were patched on the backs of hairless mice. Transepidermal water loss was measured to evaluate the skin barrier dysfunction caused by the proteases. Papain or that treated with an irreversible inhibitor specific to cysteine proteases, E64, was painted onto the ear lobes of mice of an inbred strain C57BL/6. Serum total IgE levels and papain-specific IgE and IgG antibodies were measured by ELISA.
RESULTS
Papain and V8 protease patched on the backs of hairless mice caused skin barrier dysfunction and increased serum total IgE levels, and papain induced the production of papain-specific IgG1, IgG2a, and IgG2b. Papain painted onto the ear lobes of C57BL/6 mice induced papain-specific IgE, IgG1, IgG2c, and IgG2b, whereas papain treated with E64 did not. IgG1 was the most significantly induced papain-specific IgG subclass among those measured.
CONCLUSIONS
We demonstrated that the epicutaneous administration of protease not only disrupted skin barrier function, but also induced IgE and IgG responses in a manner dependent on its protease activity. These results suggest that protease activity contained in environmental sources contributes to sensitization through an epicutaneous route.
Topics: Allergens; Animals; Antibody Specificity; Cysteine Proteases; Dermatitis, Atopic; Disease Models, Animal; Female; Hypersensitivity, Immediate; Immunoglobulin E; Immunoglobulin G; Mice; Papain; Skin
PubMed: 24662805
DOI: 10.2332/allergolint.13-OA-0621 -
BMC Genomics Jan 2018Papain-like cysteine proteases (PLCPs), a large group of cysteine proteases structurally related to papain, play important roles in plant development, senescence, and...
BACKGROUND
Papain-like cysteine proteases (PLCPs), a large group of cysteine proteases structurally related to papain, play important roles in plant development, senescence, and defense responses. Papain, the first cysteine protease whose structure was determined by X-ray crystallography, plays a crucial role in protecting papaya from herbivorous insects. Except the four major PLCPs purified and characterized in papaya latex, the rest of the PLCPs in papaya genome are largely unknown.
RESULTS
We identified 33 PLCP genes in papaya genome. Phylogenetic analysis clearly separated plant PLCP genes into nine subfamilies. PLCP genes are not equally distributed among the nine subfamilies and the number of PLCPs in each subfamily does not increase or decrease proportionally among the seven selected plant species. Papaya showed clear lineage-specific gene expansion in the subfamily III. Interestingly, all four major PLCPs purified from papaya latex, including papain, chymopapain, glycyl endopeptidase and caricain, were grouped into the lineage-specific expansion branch in the subfamily III. Mapping PLCP genes on chromosomes of five plant species revealed that lineage-specific expansions of PLCP genes were mostly derived from tandem duplications. We estimated divergence time of papaya PLCP genes of subfamily III. The major duplication events leading to lineage-specific expansion of papaya PLCP genes in subfamily III were estimated at 48 MYA, 34 MYA, and 16 MYA. The gene expression patterns of the papaya PLCP genes in different tissues were assessed by transcriptome sequencing and qRT-PCR. Most of the papaya PLCP genes of subfamily III expressed at high levels in leaf and green fruit tissues.
CONCLUSIONS
Tandem duplications played the dominant role in affecting copy number of PLCPs in plants. Significant variations in size of the PLCP subfamilies among species may reflect genetic adaptation of plant species to different environments. The lineage-specific expansion of papaya PLCPs of subfamily III might have been promoted by the continuous reciprocal selective effects of herbivore attack and plant defense.
Topics: Carica; Cell Lineage; Gene Duplication; Genome, Plant; High-Throughput Nucleotide Sequencing; Multigene Family; Papain; Phylogeny; Plant Proteins
PubMed: 29306330
DOI: 10.1186/s12864-017-4394-y -
Journal of Biomolecular Structure &... Apr 2022Novel coronavirus SARS-CoV-2 has infected 18 million people with 700,000+ mortalities worldwide and this deadly numeric figure is rapidly rising. With very few success...
Novel coronavirus SARS-CoV-2 has infected 18 million people with 700,000+ mortalities worldwide and this deadly numeric figure is rapidly rising. With very few success stories, the therapeutic targeting of this epidemic has been mainly attributed to main protease (Mpro), whilst Papain-like proteases (PLpro) also plays a vital role in the processing of replicase polyprotein. Multifunctional roles of PLpro such as viral polypeptide cleavage, de-ISGlyation and immune suppression have made it a promising drug target for therapeutic interventions. Whilst there have been a number of studies and others are on-going on repurposing and new-small molecule screening, albeit previously FDA approved drugs viz. Chloroquine (CQ) and Hydroxychloroquine (HCQ) have only been found effective against this pandemic. Inspired by this fact, we have carried out molecular docking and dynamics simulation studies of FDA approved CQ and HCQ against SARS-CoV-2 PLpro. The end aim is to characterise the binding mode of CQ and HCQ and identify the key amino acid residues involved in the mechanism of action. Further, molecular dynamics simulations (MDS) were carried out with the docked complex to search for the conformational space and for understanding the integrity of binding mode. We showed that the CQ and HCQ can bind with better binding affinity with PLpro as compared to reference known PLpro inhibitor. Based on the presented findings, it can be anticipated that the SARS-CoV-2 PLpro may act as molecular target of CQ and HCQ, and can be projected for further exploration to design potent inhibitors of SARS-CoV-2 PLpro in the near future.
Topics: Antiviral Agents; Chloroquine; Coronavirus Papain-Like Proteases; Humans; Hydroxychloroquine; Molecular Docking Simulation; Molecular Dynamics Simulation; Papain; SARS-CoV-2; COVID-19 Drug Treatment
PubMed: 33200683
DOI: 10.1080/07391102.2020.1844804 -
International Journal of Molecular... Sep 2022In late 2019, a new coronavirus (CoV) caused the outbreak of a deadly respiratory disease, resulting in the COVID-19 pandemic. In view of the ongoing pandemic, there is...
In late 2019, a new coronavirus (CoV) caused the outbreak of a deadly respiratory disease, resulting in the COVID-19 pandemic. In view of the ongoing pandemic, there is an immediate need to find drugs to treat patients. SARS-CoV-2 papain-like cysteine protease (PLpro) not only plays an important role in the pathogenesis of the virus but is also a target protein for the development of inhibitor drugs. Therefore, to develop targeted inhibitors, it is necessary to analyse and verify PLpro sites and explore whether there are other cryptic binding pockets with better activity. In this study, first, we detected the site of the whole PLpro protein by sitemap of Schrödinger (version 2018), the cavity of LigBuilder V3, and DeepSite, and roughly judged the possible activated binding site area. Then, we used the mixed solvent dynamics simulation (MixMD) of probe molecules to induce conformational changes in the protein to find the possible cryptic active sites. Finally, the TRAPP method was used to predict the druggability of cryptic pockets and analyse the changes in the physicochemical properties of residues around these sites. This work will help promote the research of SARS-CoV-2 PLpro inhibitors.
Topics: Amino Acid Sequence; Coronavirus Papain-Like Proteases; Humans; Pandemics; Papain; SARS-CoV-2; Solvents; COVID-19 Drug Treatment
PubMed: 36232570
DOI: 10.3390/ijms231911265 -
Antiviral Research Oct 2018Porcine epidemic diarrhea virus (PEDV) is a coronavirus (CoV) discovered in the 1970s that infects the intestinal tract of pigs, resulting in diarrhea and vomiting. It...
Porcine epidemic diarrhea virus (PEDV) is a coronavirus (CoV) discovered in the 1970s that infects the intestinal tract of pigs, resulting in diarrhea and vomiting. It can cause extreme dehydration and death in neonatal piglets. In Asia, modified live attenuated vaccines have been used to control PEDV infection in recent years. However, a new strain of PEDV that belongs to genogroup 2a appeared in the USA in 2013 and then quickly spread to Canada and Mexico as well as Asian and European countries. Due to the less effective protective immunity provided by the vaccines against this new strain, it has caused considerable agricultural and economic loss worldwide. The emergence of this new strain increases the importance of understanding PEDV as well as strategies for inhibiting it. Coronaviral proteases, including main proteases and papain-like proteases, are ideal antiviral targets because of their essential roles in viral maturation. Here we provide a first description of the expression, purification and structural characteristics of recombinant PEDV papain-like protease 2, moreover present our finding that 6-thioguanine, a chemotherapeutic drug, in contrast to its competitive inhibition on SARS- and MERS-CoV papain-like proteases, is a noncompetitive inhibitor of PEDV papain-like protease 2.
Topics: Antiviral Agents; Binding Sites; Coronavirus; Coronavirus Infections; Coronavirus Papain-Like Proteases; Kinetics; Molecular Docking Simulation; Papain; Porcine epidemic diarrhea virus; Protein Conformation; Recombinant Proteins; Thioguanine
PubMed: 30138642
DOI: 10.1016/j.antiviral.2018.08.011 -
Parasites & Vectors Jun 2021Plant-derived cysteine proteinases of the papain family (CPs) attack nematodes by digesting the cuticle, leading to rupture and death of the worm. The nematode cuticle...
BACKGROUND
Plant-derived cysteine proteinases of the papain family (CPs) attack nematodes by digesting the cuticle, leading to rupture and death of the worm. The nematode cuticle is composed of collagens and cuticlins, but the specific molecular target(s) for the proteinases have yet to be identified.
METHODS
This study followed the course of nematode cuticle disruption using immunohistochemistry, scanning electron microscopy and proteomics, using a free-living nematode, Caenorhabditis elegans and the murine GI nematode Heligmosomoides bakeri (H. polygyrus) as target organisms.
RESULTS
Immunohistochemistry indicated that DPY-7 collagen is a target for CPs on the cuticle of C. elegans. The time course of loss of DPY-7 from the cuticle allowed us to use it to visualise the process of cuticle disruption. There was a marked difference in the time course of damage to the cuticles of the two species of nematode, with H. bakeri being more rapidly hydrolysed. In general, the CPs' mode of attack on the nematode cuticle was by degrading the structural proteins, leading to loss of integrity of the cuticle, and finally death of the nematode. Proteomic analysis failed conclusively to identify structural targets for CPs, but preliminary data suggested that COL-87 and CUT-19 may be important targets for the CPs, the digestion of which may contribute to cuticle disruption and death of the worm. Cuticle globin was also identified as a cuticular target. The presence of more than one target protein may slow the development of resistance against this new class of anthelmintic.
CONCLUSIONS
Scanning electron microscopy and immunohistochemistry allowed the process of disruption of the cuticle to be followed with time. Cuticle collagens and cuticlins are molecular targets for plant cysteine proteinases. However, the presence of tyrosine cross-links in nematode cuticle proteins seriously impeded protein identification by proteomic analyses. Multiple cuticle targets exist, probably making resistance to this new anthelmintic slow to develop.
Topics: Animals; Anthelmintics; Caenorhabditis elegans; Cysteine Proteases; Female; Male; Mice; Nematoda; Papain; Plant Extracts; Proteomics
PubMed: 34090505
DOI: 10.1186/s13071-021-04800-8 -
PLoS Pathogens Dec 2023Deubiquitination of cellular substrates by viral proteases is a mechanism used to interfere with host cellular signaling processes, shared between members of the...
Demonstrating the importance of porcine reproductive and respiratory syndrome virus papain-like protease 2 deubiquitinating activity in viral replication by structure-guided mutagenesis.
Deubiquitination of cellular substrates by viral proteases is a mechanism used to interfere with host cellular signaling processes, shared between members of the coronavirus- and arterivirus families. In the case of Arteriviruses, deubiquitinating and polyprotein processing activities are accomplished by the virus-encoded papain-like protease 2 (PLP2). Several studies have implicated the deubiquitinating activity of the porcine reproductive and respiratory syndrome virus (PRRSV) PLP2 in the downregulation of cellular interferon production, however to date, the only arterivirus PLP2 structure described is that of equine arteritis virus (EAV), a distantly related virus. Here we describe the first crystal structure of the PRRSV PLP2 domain both in the presence and absence of its ubiquitin substrate, which reveals unique structural differences in this viral domain compared to PLP2 from EAV. To probe the role of PRRSV PLP2 deubiquitinating activity in host immune evasion, we selectively removed this activity from the domain by mutagenesis and found that the viral domain could no longer downregulate cellular interferon production. Interestingly, unlike EAV, and also unlike the situation for MERS-CoV, we found that recombinant PRRSV carrying PLP2 DUB-specific mutations faces significant selective pressure to revert to wild-type virus in MARC-145 cells, suggesting that the PLP2 DUB activity, which in PRRSV is present as three different versions of viral protein nsp2 expressed during infection, is critically important for PRRSV replication.
Topics: Animals; Horses; Swine; Humans; Papain; Porcine respiratory and reproductive syndrome virus; Mutagenesis; Equartevirus; Peptide Hydrolases; Virus Replication; Interferons; Viral Nonstructural Proteins
PubMed: 38096325
DOI: 10.1371/journal.ppat.1011872 -
Scientific Reports Oct 2022The interest in agri-food residues and their valorization has grown considerably, and many of them are today considered to be valuable, under-exploited sources of...
The interest in agri-food residues and their valorization has grown considerably, and many of them are today considered to be valuable, under-exploited sources of different compounds and notably proteins. Despite the beneficial properties of legumes by-products, there are also some emerging risks to consider, including their potential allergenicity. In this work the immunoreactivity of chickpea, pea, and white bean by-products was assessed, and whether the production of enzymatic hydrolysates can be an effective strategy to reduce this allergenic potential. The results presented clearly indicate that the efficiency of this strategy is strongly related to the enzyme used and the food matrix. All legume by-products showed immunoreactivity towards serum of legume-allergic patients. Hydrolysates from alcalase did not show residual immunoreactivity for chickpea and green pea, whereas hydrolysates from papain still presented some immunoreactivity. However, for white beans, the presence of antinutritional factors prevented a complete hydrolysis, yielding a residual immunoreactivity even after enzymatic hydrolysis with alcalase.
Topics: Allergens; Cicer; Fabaceae; Humans; Hydrolysis; Papain; Protein Hydrolysates; Subtilisins
PubMed: 36207409
DOI: 10.1038/s41598-022-21296-z -
International Journal of Molecular... Jul 2023This study investigates the features of interactions between cysteine proteases (bromelain, ficin, and papain) and a graft copolymer of carboxymethyl cellulose sodium...
This study investigates the features of interactions between cysteine proteases (bromelain, ficin, and papain) and a graft copolymer of carboxymethyl cellulose sodium salt with -vinylimidazole. The objective is to understand the influence of this interactions on the proteolytic activity and stability of the enzymes. The enzymes were immobilized through complexation with the carrier. The interaction mechanism was examined using Fourier-transform infrared spectroscopy and flexible molecular docking simulations. The findings reveal that the enzymes interact with the functional groups of the carrier via amino acid residues, resulting in the formation of secondary structure elements and enzyme's active sites. These interactions induce modulation of active site of the enzymes, leading to an enhancement in their proteolytic activity. Furthermore, the immobilized enzymes demonstrate superior stability compared to their native counterparts. Notably, during a 21-day incubation period, no protein release from the conjugates was observed. These results suggest that the complexation of the enzymes with the graft copolymer has the potential to improve their performance as biocatalysts, with applications in various fields such as biomedicine, pharmaceutics, and biotechnology.
Topics: Papain; Bromelains; Ficain; Carboxymethylcellulose Sodium; Molecular Docking Simulation; Polymers; Sodium Chloride; Sodium Chloride, Dietary; Sodium
PubMed: 37511006
DOI: 10.3390/ijms241411246