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Immunity Apr 2020Innate lymphoid cells (ILCs) play an important role in the control and maintenance of barrier immunity. However, chronic activation of ILCs results in immune-mediated...
Innate lymphoid cells (ILCs) play an important role in the control and maintenance of barrier immunity. However, chronic activation of ILCs results in immune-mediated pathology. Here, we show that tissue-resident type 2 ILCs (ILC2s) display a distinct metabolic signature upon chronic activation. In the context of allergen-driven airway inflammation, ILC2s increase their uptake of both external lipids and glucose. Externally acquired fatty acids are transiently stored in lipid droplets and converted into phospholipids to promote the proliferation of ILC2s. This metabolic program is imprinted by interleukin-33 (IL-33) and regulated by the genes Pparg and Dgat1, which are both controlled by glucose availability and mTOR signaling. Restricting dietary glucose by feeding mice a ketogenic diet largely ablated ILC2-mediated airway inflammation by impairing fatty acid metabolism and the formation of lipid droplets. Together, these results reveal that pathogenic ILC2 responses require lipid metabolism and identify ketogenic diet as a potent intervention strategy to treat airway inflammation.
Topics: Allergens; Alternaria; Animals; Asthma; Cell Lineage; Cytokines; Diacylglycerol O-Acyltransferase; Diet, Ketogenic; Disease Models, Animal; Fatty Acids; Gene Expression Regulation; Glucose; Immunity, Innate; Interleukin-33; Interleukins; Lipid Droplets; Lung; Mice; Mice, Inbred C57BL; Mice, Knockout; PPAR gamma; Papain; Phospholipids; Primary Cell Culture; T-Lymphocyte Subsets; TOR Serine-Threonine Kinases; Thymic Stromal Lymphopoietin
PubMed: 32268121
DOI: 10.1016/j.immuni.2020.03.003 -
Nature Microbiology Apr 2021Activation of the RIG-I-like receptors, retinoic-acid inducible gene I (RIG-I) and melanoma differentiation-associated protein 5 (MDA5), establishes an antiviral state...
Activation of the RIG-I-like receptors, retinoic-acid inducible gene I (RIG-I) and melanoma differentiation-associated protein 5 (MDA5), establishes an antiviral state by upregulating interferon (IFN)-stimulated genes (ISGs). Among these is ISG15, the mechanistic roles of which in innate immunity still remain enigmatic. In the present study, we report that ISG15 conjugation is essential for antiviral IFN responses mediated by the viral RNA sensor MDA5. ISGylation of the caspase activation and recruitment domains of MDA5 promotes its oligomerization and thereby triggers activation of innate immunity against a range of viruses, including coronaviruses, flaviviruses and picornaviruses. The ISG15-dependent activation of MDA5 is antagonized through direct de-ISGylation mediated by the papain-like protease of SARS-CoV-2, a recently emerged coronavirus that has caused the COVID-19 pandemic. Our work demonstrates a crucial role for ISG15 in the MDA5-mediated antiviral response, and also identifies a key immune evasion mechanism of SARS-CoV-2, which may be targeted for the development of new antivirals and vaccines to combat COVID-19.
Topics: Aedes; Animals; Chlorocebus aethiops; Coronavirus Papain-Like Proteases; Cricetinae; Cytokines; HEK293 Cells; Humans; Immunity, Innate; Interferon-Induced Helicase, IFIH1; Leukocytes, Mononuclear; Mice; SARS-CoV-2; Ubiquitins; Vero Cells
PubMed: 33727702
DOI: 10.1038/s41564-021-00884-1 -
Chembiochem : a European Journal of... Oct 2022Emerging variants of SARS-CoV-2 and potential novel epidemic coronaviruses underline the importance of investigating various viral proteins as potential drug targets.... (Review)
Review
Emerging variants of SARS-CoV-2 and potential novel epidemic coronaviruses underline the importance of investigating various viral proteins as potential drug targets. The papain-like protease of coronaviruses has been less explored than other viral proteins; however, its substantive role in viral replication and impact on the host immune response make it a suitable target to study. This review article focuses on the structure and function of the papain-like protease (PL ) of SARS-CoV-2, including variants of concern, and compares it to those of other coronaviruses, such as SARS-CoV-1 and MERS-CoV. The protease's recognition motif is mirrored in ubiquitin and ISG15, which are involved in the antiviral immune response. Inhibitors, including GRL0617 derivatives, and their prospects as potential future antiviral agents are also discussed.
Topics: Aniline Compounds; Antiviral Agents; Benzamides; Coronavirus Papain-Like Proteases; Humans; Naphthalenes; Papain; Peptide Hydrolases; Protease Inhibitors; SARS-CoV-2; Ubiquitin; Viral Proteins; COVID-19 Drug Treatment
PubMed: 35993805
DOI: 10.1002/cbic.202200327 -
Journal of Medicinal Chemistry Jun 2022SARS-CoV-2 is the causative agent of the COVID-19 pandemic. The approval of vaccines and small-molecule antivirals is vital in combating the pandemic. The viral... (Review)
Review
SARS-CoV-2 is the causative agent of the COVID-19 pandemic. The approval of vaccines and small-molecule antivirals is vital in combating the pandemic. The viral polymerase inhibitors remdesivir and molnupiravir and the viral main protease inhibitor nirmatrelvir/ritonavir have been approved by the U.S. FDA. However, the emergence of variants of concern/interest calls for additional antivirals with novel mechanisms of action. The SARS-CoV-2 papain-like protease (PL) mediates the cleavage of viral polyprotein and modulates the host's innate immune response upon viral infection, rendering it a promising antiviral drug target. This Perspective highlights major achievements in structure-based design and high-throughput screening of SARS-CoV-2 PL inhibitors since the beginning of the pandemic. Encouraging progress includes the design of non-covalent PL inhibitors with favorable pharmacokinetic properties and the first-in-class covalent PL inhibitors. In addition, we offer our opinion on the knowledge gaps that need to be filled to advance PL inhibitors to the clinic.
Topics: Antiviral Agents; Coronavirus Papain-Like Proteases; Humans; Pandemics; Protease Inhibitors; SARS-CoV-2; COVID-19 Drug Treatment
PubMed: 35620927
DOI: 10.1021/acs.jmedchem.2c00303 -
International Journal of Biological... Oct 2021Papain is a cysteine protease from papaya, with many applications due to its broad specificity. This paper reviews for first time the immobilization of papain on... (Review)
Review
Papain is a cysteine protease from papaya, with many applications due to its broad specificity. This paper reviews for first time the immobilization of papain on different supports (organic, inorganic or hybrid supports) presenting some of the features of the utilized immobilization strategies (e.g., epoxide, glutaraldehyde, genipin, glyoxyl for covalent immobilization). Special focus is placed on the preparation of magnetic biocatalysts, which will permit the simple recovery of the biocatalyst even if the medium is a suspension. Problems specific to the immobilization of proteases (e.g., steric problems when hydrolyzing large proteins) are also defined. The benefits of a proper immobilization (enzyme stabilization, widening of the operation window) are discussed, together with some artifacts that may suggest an enzyme stabilization that may be unrelated to enzyme rigidification.
Topics: Carica; Enzyme Stability; Enzymes; Enzymes, Immobilized; Papain
PubMed: 34375660
DOI: 10.1016/j.ijbiomac.2021.08.016 -
FASEB Journal : Official Publication of... Jan 2021SARS-CoV and SARS-CoV-2 encode four structural and accessory proteins (spike, envelope, membrane and nucleocapsid proteins) and two polyproteins (pp1a and pp1ab). The... (Review)
Review
SARS-CoV and SARS-CoV-2 encode four structural and accessory proteins (spike, envelope, membrane and nucleocapsid proteins) and two polyproteins (pp1a and pp1ab). The polyproteins are further cleaved by 3C-like cysteine protease (3CL ) and papain-like protease (PL ) into 16 nonstructural proteins (nsps). PL is released from nsp3 through autocleavage, and then it cleaves the sites between nsp1/2, between nsp2/3 and between nsp3/4 with recognition motif of LXGG, and the sites in the C-terminus of ubiquitin and of protein interferon-stimulated gene 15 (ISG15) with recognition motif of RLRGG. Alone or together with SARS unique domain (SUD), PL can stabilize an E3 ubiquitin ligase, the ring-finger, and CHY zinc-finger domain-containing 1 (RCHY1), through domain interaction, and thus, promote RCHY1 to ubiquitinate its target proteins including p53. However, a dilemma appears in terms of PL roles. On the one hand, the ubiquitination of p53 is good for SARS-CoV because the ubiquitinated p53 cannot inhibit SARS-CoV replication. On the other hand, the ubiquitination of NF-κB inhibitor (IκBα), TNF receptor-associated factors (TRAFs), and stimulator of interferon gene (STING), and the ISGylation of targeted proteins are bad for SARS-CoV because these ubiquitination and ISGylation initiate the innate immune response and antiviral state. This mini-review analyzes the dilemma and provides a snapshot on how the viral PL smartly manages its roles to avoid its simultaneously contradictory actions, which could shed lights on possible strategies to deal with SARS-CoV-2 infections.
Topics: COVID-19; Coronavirus Papain-Like Proteases; Genes, Viral; Host-Pathogen Interactions; Humans; Molecular Targeted Therapy; NF-kappa B; Protein Domains; Protein Processing, Post-Translational; Severe acute respiratory syndrome-related coronavirus; SARS-CoV-2; Severe Acute Respiratory Syndrome; Substrate Specificity; Ubiquitin-Activating Enzymes; Ubiquitin-Conjugating Enzymes; Ubiquitin-Protein Ligases; Ubiquitination; Viral Proteins; Virus Replication
PubMed: 33368679
DOI: 10.1096/fj.202002271 -
The Journal of Biological Chemistry Jun 2023Papain-like cysteine peptidases form a big and highly diverse superfamily of proteins involved in many important biological functions, such as protein turnover,...
Papain-like cysteine peptidases form a big and highly diverse superfamily of proteins involved in many important biological functions, such as protein turnover, deubiquitination, tissue remodeling, blood clotting, virulence, defense, and cell wall remodeling. High sequence and structure diversity observed within these proteins hinders their comprehensive classification as well as the identification of new representatives. Moreover, in general protein databases, many families already classified as papain like lack details regarding their mechanism of action or biological function. Here, we use transitive remote homology searches and 3D modeling to newly classify 21 families to the papain-like cysteine peptidase superfamily. We attempt to predict their biological function and provide structural characterization of 89 protein clusters defined based on sequence similarity altogether spanning 106 papain-like families. Moreover, we systematically discuss observed diversity in sequences, structures, and catalytic sites. Eventually, we expand the list of human papain-related proteins by seven representatives, including dopamine receptor-interacting protein 1 as potential deubiquitinase, and centriole duplication regulating CEP76 as retaining catalytically active peptidase-like domain. The presented results not only provide structure-based rationales to already existing peptidase databases but also may inspire further experimental research focused on peptidase-related biological processes.
Topics: Humans; Catalytic Domain; Centrioles; Cysteine Proteases; Deubiquitinating Enzymes; Models, Molecular; Papain; Databases, Protein
PubMed: 37164157
DOI: 10.1016/j.jbc.2023.104801 -
Scientific Reports Sep 2021In this pandemic SARS-CoV-2 crisis, any attempt to contain and eliminate the virus will also stop its spread and consequently decrease the risk of severe illness and...
In this pandemic SARS-CoV-2 crisis, any attempt to contain and eliminate the virus will also stop its spread and consequently decrease the risk of severe illness and death. While ozone treatment has been suggested as an effective disinfection process, no precise mechanism of action has been previously reported. This study aimed to further investigate the effect of ozone treatment on SARS-CoV-2. Therefore, virus collected from nasopharyngeal and oropharyngeal swab and sputum samples from symptomatic patients was exposed to ozone for different exposure times. The virus morphology and structure were monitored and analyzed through Atomic Force Microscopy (AFM), Transmission Electron Microscopy (TEM), Atomic Absorption Spectroscopy (AAS), and ATR-FTIR. The obtained results showed that ozone treatment not only unsettles the virus morphology but also alters the virus proteins' structure and conformation through amino acid disturbance and Zn ion release from the virus non-structural proteins. These results could provide a clearer pathway for virus elimination and therapeutics preparation.
Topics: Coronavirus Papain-Like Proteases; Coronavirus RNA-Dependent RNA Polymerase; Humans; Microscopy, Electron, Transmission; Ozone; Protein Structure, Secondary; Protein Structure, Tertiary; SARS-CoV-2; Time Factors; Viral Envelope; Viral Regulatory and Accessory Proteins; Zinc; COVID-19 Drug Treatment
PubMed: 34552128
DOI: 10.1038/s41598-021-97860-w -
Ibrain 2022The objective of this study was to compare the efficiency of trypsin and papain in neuronal digestion and determine which enzyme is more efficient. Cortical tissues were...
The objective of this study was to compare the efficiency of trypsin and papain in neuronal digestion and determine which enzyme is more efficient. Cortical tissues were obtained from Sprague-Dawley (SD) rats. According to the different digestive enzymes, the samples were divided into the trypsin group and the papain group. After being digested by each of the two enzymes, cortical neurons were collected from the samples. Then, the morphology of the cortical neurons was determined. Moreover, the cortical neurons were transfected with the negative control (NC) lentivirus. The transfection efficiency and morphology were determined and compared. Compared with the papain group, cortical neurons in the trypsin group were more in number, had larger cell size, had longer axonal length, and had fewer impurities. The transfection efficiency of the trypsin group (57.77%) was higher than that of the papain group (53.83%). The morphology of neurons that was displayed showed that the cell body of most neurons shrank and became smaller, and the axis mutation became shorter and less in the papain group 6 days after transfection with the NC lentivirus. Trypsin is more efficient in digesting neurons because the neurons digested by this enzyme are more in number, have a larger cell body, longer axons, and greater transfection efficiency.
PubMed: 37786412
DOI: 10.1002/ibra.12028 -
PloS One 2023Hypertrophic scars and keloids are characterized by an excessive collagen deposition. The available treatment options are invasive and can result in recurrence of scar...
Hypertrophic scars and keloids are characterized by an excessive collagen deposition. The available treatment options are invasive and can result in recurrence of scar formation. Using liposomes and transferosomes for the topical delivery of papain, a proteolytic enzyme, can be effective treatment. The objective of the study is to formulate papain-loaded liposomes and transferosomes, characterize the formulations, and study in vitro permeation using shed snake skin and Sprague-Dawley rat skin as models for stratum corneum and full thickness skin. Papain-loaded liposomes and transferosomes were formulated using the thin-film hydration method for the delivery of papain across the stratum corneum barrier. An in vitro permeation study carried out using shed-snake skin and Sprague-Dawley rat skin models showed that transferosomes were able to deliver papain across the stratum corneum barrier, while papain solution and papain liposomes were not able to cross the barrier. However, transferosomes were not able to deliver papain across the full thickness rat skin model suggesting the deposition of papain loaded transferosomes in the epidermal or dermal layer of skin. In addition, an ex-vivo model was used to analyze the effect of papain exposure on the morphology of the epidermis taken from rat skin exposed to papain solution, papain in transferosomes and papain in liposomes. Papain in solution resulted in a noticeable degradation of the epidermis, but when embedded in either transferosomes or liposomes there was no noticeable change when compared to control animals. The cytotoxicity study performed using HeLa cells showed that the cells were viable at papain concentrations lower than 0.01 mg/ml.
Topics: Humans; Rats; Animals; Liposomes; Keloid; Cicatrix, Hypertrophic; Papain; Rats, Sprague-Dawley; HeLa Cells; Administration, Cutaneous
PubMed: 38100466
DOI: 10.1371/journal.pone.0290224