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IUBMB Life Mar 2019The plant toxin, abrin, a type-II ribosome inactivating protein, is extremely lethal, the human fatal dose being ~1 μg/kg body weight. Abrin has been classified as an...
The plant toxin, abrin, a type-II ribosome inactivating protein, is extremely lethal, the human fatal dose being ~1 μg/kg body weight. Abrin has been classified as an agent for bioterrorism, which is of concern. Conversely, the high toxic property of abrin has been employed in generating immunotoxins, whereas its toxin moiety is conjugated to cell surface marker-specific antibodies for cell-targeted killing. Different cell types exhibit variable levels of sensitivity to abrin toxicity; therefore, adequate knowledge of the molecular mechanism that governs the activity of the protein would be a safeguard. To gain insights into this, two cell lines requiring strikingly different concentrations of abrin for inactivating ribosomes were studied. Employing conjugates of the wild-type and active site mutant of abrin A chain with the ricin B chain, it was found that abrin-induced apoptosis was dependent on inhibition of protein synthesis (PSI) leading to ER-stress in Ovcar-3 cells, but not in KB cells. Abrin was also observed to cause direct DNA damage in KB cells, while in Ovcar-3 cells abrin-induced DNA damage was found to be dependent on caspases. Overall, the study demonstrates that the correlation of abrin-mediated PSI and apoptosis is cell-specific and abrin can induce more than one pathway to cause cell death. © 2018 IUBMB Life, 71(3):357-363, 2019.
Topics: Abrin; Apoptosis; Cell Line, Tumor; DNA Damage; Dose-Response Relationship, Drug; Endoplasmic Reticulum Stress; Glycoconjugates; HeLa Cells; Humans; Mutation; Organ Specificity; Protein Biosynthesis; Recombinant Proteins; Ribosomes; Ricin; Structure-Activity Relationship
PubMed: 30506989
DOI: 10.1002/iub.1980 -
Toxins May 2021The high toxic abrin from the plant is a type II ribosome-inactivating protein toxin with a human lethal dose of 0.1-1.0 µg/kg body weight. Due to its high toxicity...
The high toxic abrin from the plant is a type II ribosome-inactivating protein toxin with a human lethal dose of 0.1-1.0 µg/kg body weight. Due to its high toxicity and the potential misuse as a biothreat agent, it is of great importance to developing fast and reliable methods for the identification and quantification of abrin in complex matrices. Here, we report rapid and efficient acetonitrile (ACN)- and ultrasound-assisted on-bead trypsin digestion method combined with HPLC-MS/MS for the quantification of abrin isoforms in complex matrices. Specific peptides of abrin isoforms were generated by direct ACN-assisted trypsin digestion and analyzed by HPLC-HRMS. Combined with in silico digestion and BLASTp database search, fifteen marker peptides were selected for differential detection of abrin isoforms. The abrin in milk and plasma was enriched by immunomagnetic beads prepared by biotinylated anti-abrin polyclonal antibodies conjugated to streptavidin magnetic beads. The ultrasound-assisted on-bead trypsin digestion method was carried out under the condition of 10% ACN as denaturant solvent, the entire digestion time was further shortened from 90 min to 30 min. The four peptides of T3A, T12A, T15A and T9A were chosen as quantification for total abrin, abrin-a, abrin-b, and abrin-c/d, respectively. The absolute quantification of abrin and its isoforms was accomplished by isotope dilution with labeled AQUA peptides and analyzed by HPLC-MS/MS (MRM). The developed method was fully validated in milk and plasma matrices with quantification limits in the range of 1.0-9.4 ng/mL for the isoforms of abrin. Furthermore, the developed approach was applied for the characterization of abrin isoforms from various fractions from gel filtration separation of the seeds, and measurement of abrin in the samples of biotoxin exercises organized by the Organization for the Prohibition of Chemical Weapons (OPCW). This study provided a recommended method for the differential identification of abrin isoforms, which are easily applied in international laboratories to improve the capabilities for the analysis of biotoxin samples.
Topics: Abrin; Abrus; Animals; Chromatography, High Pressure Liquid; Chromatography, Liquid; Computer Simulation; Milk; Protein Isoforms; Rabbits; Tandem Mass Spectrometry; Toxins, Biological; Trypsin; Ultrasonics
PubMed: 34069935
DOI: 10.3390/toxins13050358 -
Toxins Feb 2022Abrin is a types II ribosome-inactivating protein (RIP) isolated from seeds, which comprises a catalytically active A chain and a lectin-like B chain linked by a...
Abrin is a types II ribosome-inactivating protein (RIP) isolated from seeds, which comprises a catalytically active A chain and a lectin-like B chain linked by a disulfide bond. Four isotoxins of abrin have been reported with similar amino-acid composition but different cytotoxicity, of which abrin-a is the most potent toxin. High lethality and easy availability make abrin a potential bioterrorism agent. However, there are no antidotes available for managing abrin poisoning, and treatment is only symptomatic. Currently, neutralizing antibodies remain the most effective therapy against biotoxin poisoning. In this study, we prepared, identified, and acquired a high-affinity neutralizing monoclonal antibody (mAb) 10D8 with a potent pre- and post-exposure protective effect against cytotoxicity and animal toxicity induced by abrin-a or abrin crude extract. The mAb 10D8 could rescue the mouse injected intraperitoneally with a 25 × LD dose of abrin-a from lethality and prevent tissue damages. Results indicated that 10D8 does not prevent the binding and internalization of abrin-a to cells but inhibits the enzymatic activity of abrin-a and reduces protein synthesis inhibition of cells. The high affinity, good specificity, and potent antitoxic efficiency of 10D8 make it a promising candidate for therapeutic antibodies against abrin.
Topics: Abrin; Abrus; Animals; Antibodies, Monoclonal; Antibodies, Neutralizing; Antitoxins; Mice
PubMed: 35324661
DOI: 10.3390/toxins14030164 -
Journal of Food Protection Sep 2008The oral and intraperitoneal (i.p.) toxicities to female BALB/c mice of ricin and abrin in phosphate-buffered saline (PBS), spring water, apple juice, and half-and-half...
The oral and intraperitoneal (i.p.) toxicities to female BALB/c mice of ricin and abrin in phosphate-buffered saline (PBS), spring water, apple juice, and half-and-half (only oral) were examined after brief (2 h) and prolonged (11 to 13 days) storage. The ricin and abrin samples prepared in PBS had oral toxicities consistent with those previous studies, indicating oral and i.p. 50% lethal doses of > 1 mg/kg of body weight and between 2 and 20 microg/kg of body weight, respectively. The toxicities of ricin and abrin in PBS were greater than those in apple juice and water. The oral toxicity of ricin and abrin in half-and-half appeared comparable to or less than that observed for the toxins in water. Spiked samples stored for a maximum of 11 days (13 for the abrin samples) at 4 degrees C induced similar numbers of fatalities as did samples stored for only 2 h. Enzyme-linked immunosorbent assays of the samples administered by i.p. injection indicated a decrease in detectable toxin at 0.5 microg/ml.
Topics: Abrin; Administration, Oral; Animals; Beverages; Biological Assay; Consumer Product Safety; Dose-Response Relationship, Drug; Female; Food Contamination; Humans; Injections, Intraperitoneal; Lethal Dose 50; Malus; Mice; Mice, Inbred BALB C; Milk; Random Allocation; Ricin; Temperature; Time Factors
PubMed: 18810872
DOI: 10.4315/0362-028x-71.9.1875 -
Toxins Feb 2018Biological toxins are a heterogeneous group of compounds that share commonalities with biological and chemical agents. Among them, protein toxins represent a... (Review)
Review
Biological toxins are a heterogeneous group of compounds that share commonalities with biological and chemical agents. Among them, protein toxins represent a considerable, diverse set. They cover a broad range of molecular weights from less than 1000 Da to more than 150 kDa. This review aims to compare conventional detection methods of protein toxins such as in vitro bioassays with proteomic methods, including immunoassays and mass spectrometry-based techniques and their combination. Special emphasis is given to toxins falling into a group of selected agents, according to the Centers for Disease Control and Prevention, such as , toxins, toxins, epsilon toxin, ricin from , Abrin from or control of trade in dual-use items in the European Union, including lesser known protein toxins such as Viscumin from . The analysis of protein toxins and monitoring for biological threats, i.e., the deliberate spread of infectious microorganisms or toxins through water, food, or the air, requires rapid and reliable methods for the early identification of these agents.
Topics: Animals; Bacterial Proteins; Humans; Plant Proteins; Proteomics; Toxins, Biological
PubMed: 29495560
DOI: 10.3390/toxins10030099 -
Se Pu = Chinese Journal of... Mar 2021Type Ⅱ ribosome-inactivating proteins (RIPs) are an important class of protein toxins that consist of A and B chains linked by an interchain disulfide bond. The... (Review)
Review
Type Ⅱ ribosome-inactivating proteins (RIPs) are an important class of protein toxins that consist of A and B chains linked by an interchain disulfide bond. The B-chain with lectin-like activity is responsible for binding to the galactose-containing receptors on eukaryotic cell surfaces, which is essential for A-chain internalization by endocytosis. The A-chain has -glycosidase activity that irreversibly depurinates a specific adenine from 28S ribosomal RNA (28S rRNA) and terminates protein synthesis. The synergistic effect of the A-B chain inactivates the ribosome, inhibits protein synthesis, and exhibits high cytotoxicity. Ricin and abrin that are expressed by the plants and , respectively, are typical type Ⅱ RIPs. The toxicity of ricin and abrin are 385 times and 2885 times, respectively, more that of the nerve agent VX. Owing to their ease of preparation, wide availability, and potential use as a bioterrorism agent, type Ⅱ RIPs have garnered increasing attention in recent years. Ricin is listed as a prohibited substance under schedule 1A of the Chemical Weapons Convention (CWC). The occurrence of ricin-related bioterrorism incidents in recent years has promoted the development of accurate, sensitive, and rapid detection and identification technology for type Ⅱ RIPs. Significant progress has been made in the study of toxicity mechanisms and detection methods of type Ⅱ RIPs, which primarily involve qualitative and quantitative analysis methods including immunological assays, mass spectrometry analysis methods, and toxin activity detection methods based on depurination and cytotoxicity. Immunoassays generally involve the specific recognition of antigens and antibodies, which is based on oligonucleotide molecular recognition elements called aptamers. These methods are fast and highly sensitive, but for highly homologous proteins in complex samples, they provide false positive results. With the rapid development of biological mass spectrometry detection technology, techniques such as electrospray ionization (ESI) and matrix-assisted laser desorption ionization (MALDI) are widely used in the identification of proteins. These methods not only provide accurate information on molecular weight and structure of proteins, but also demonstrate accurate quantification. Enzyme digestion combined with mass spectrometry is the predominantly used detection method. Accurate identification of protein toxins can be achieved by fingerprint analysis of enzymatically digested peptides. For analysis of protein toxins in complex samples, abundant peptide markers are obtained using a multi-enzyme digestion strategy. Targeted mass spectrometry analysis of peptide markers is used to obtain accurate qualitative and quantitative information, which effectively improves the accuracy and sensitivity of the identification of type Ⅱ RIP toxins. Although immunoassay and mass spectrometry detection methods can provide accurate identification of type Ⅱ RIPs, they cannot determine whether the toxins will retain potency. The widely used detection methods for activity analysis of type Ⅱ RIPs include depurination assay based on -glycosidase activity and cytotoxicity assay. Both the methods provide simple, rapid, and sensitive analysis of type Ⅱ RIP toxicity, and complement other detection methods. Owing to the importance of type Ⅱ RIP toxins, the Organization for the Prohibition of Chemical Weapons (OPCW) has proposed clear technical requirements for the identification and analysis of relevant samples. We herein reviewed the structural characteristics, mechanism of action, and the development and application of type Ⅱ RIP detection methods; nearly 70 studies on type Ⅱ RIP toxins and their detection methods have been cited. In addition to the technical requirements of OPCW for the unambiguous identification of biotoxins, the trend of future development of type Ⅱ RIP-based detection technology has been explored.
Topics: Abrin; Plant Proteins; Ribosome Inactivating Proteins; Ribosomes; Ricin; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
PubMed: 34227307
DOI: 10.3724/SP.J.1123.2020.10001 -
Toxins Sep 2022Abrin is a highly toxic protein obtained from the seeds of the rosary pea plant , and it is closely related to ricin in terms of its structure and chemical properties....
Abrin is a highly toxic protein obtained from the seeds of the rosary pea plant , and it is closely related to ricin in terms of its structure and chemical properties. Both toxins inhibit ribosomal function, halt protein synthesis and lead to cellular death. The major clinical manifestations following pulmonary exposure to these toxins consist of severe lung inflammation and consequent respiratory insufficiency. Despite the high similarity between abrin and ricin in terms of disease progression, the ability to protect mice against these toxins by postexposure antibody-mediated treatment differs significantly, with a markedly higher level of protection achieved against abrin intoxication. In this study, we conducted an in-depth comparison between the kinetics of in vivo abrin and ricin intoxication in a murine model. The data demonstrated differential binding of abrin and ricin to the parenchymal cells of the lungs. Accordingly, toxin-mediated injury to the nonhematopoietic compartment was shown to be markedly lower in the case of abrin intoxication. Thus, profiling of alveolar epithelial cells demonstrated that although toxin-induced damage was restricted to alveolar epithelial type II cells following abrin intoxication, as previously reported for ricin, it was less pronounced. Furthermore, unlike following ricin intoxication, no direct damage was detected in the lung endothelial cell population following abrin exposure. Reduced impairment of intercellular junction molecules following abrin intoxication was detected as well. In contrast, similar damage to the endothelial surface glycocalyx layer was observed for the two toxins. We assume that the reduced damage to the lung stroma, which maintains a higher level of tissue integrity following pulmonary exposure to abrin compared to ricin, contributes to the high efficiency of the anti-abrin antibody treatment at late time points after exposure.
Topics: Abrin; Abrus; Animals; Foodborne Diseases; Lung; Lung Injury; Mice; Plant Poisoning; Ricin; Toxins, Biological
PubMed: 36136552
DOI: 10.3390/toxins14090614 -
Toxins Feb 2018Abrin toxin is a type 2 ribosome inactivating glycoprotein isolated from the seeds of (jequirity pea). Owing to its high toxicity, relative ease of purification and...
Abrin toxin is a type 2 ribosome inactivating glycoprotein isolated from the seeds of (jequirity pea). Owing to its high toxicity, relative ease of purification and accessibility, it is considered a biological threat agent. To date, there is no effective post-exposure treatment for abrin poisoning and passive immunization remains the most effective therapy. However, the effectiveness of anti-abrin monoclonal antibodies for post-exposure therapy following abrin intoxication has not been demonstrated. The aim of this study was to isolate high affinity anti-abrin antibodies that possess potent toxin-neutralization capabilities. An immune scFv phage-display library was constructed from an abrin-immunized rabbit and a panel of antibodies (six directed against the A subunit of abrin and four against the B subunit) was isolated and expressed as scFv-Fc antibodies. By pair-wise analysis, we found that these antibodies target five distinct epitopes on the surface of abrin and that antibodies against all these sites can bind the toxin simultaneously. Several of these antibodies (namely, RB9, RB10, RB28 and RB30) conferred high protection against pulmonary intoxication of mice, when administered six hours post exposure to a lethal dose of abrin. The data presented in this study demonstrate for the first time the efficacy of monoclonal antibodies in treatment of mice after pulmonary intoxication with abrin and promote the use of these antibodies, one or several, for post-exposure treatment of abrin intoxication.
Topics: Abrin; Animals; Cell Surface Display Techniques; Female; HeLa Cells; Humans; Immunization; Mice, Inbred ICR; Neutralization Tests; Rabbits; Single-Chain Antibodies
PubMed: 29438273
DOI: 10.3390/toxins10020080 -
Toxins Dec 2018Abrin, a highly toxic plant toxin, is a potential bioterror weapon. Work from our laboratory and others have shown that abrin is highly resistant to both thermal and pH...
Abrin, a highly toxic plant toxin, is a potential bioterror weapon. Work from our laboratory and others have shown that abrin is highly resistant to both thermal and pH inactivation methods. We sought to evaluate the effectiveness of selected food processing thermal inactivation conditions against abrin in economically important food matrices (whole milk, non-fat milk, liquid egg, and ground beef). The effectiveness of toxin inactivation was measured via three different assays: (1) In vitro cell free translation (CFT) assay, (2) Vero cell culture cytotoxicity; and the in vivo mouse intraperitoneal (ip) bioassay. For both whole and non-fat milk, complete inactivation was achieved at temperatures of ≥ 80 °C for 3 min or 134 °C for 60 s, which were higher than the normal vat/batch pasteurization or the high temperature short time pasteurization (HTST). Toxin inactivation in liquid egg required temperatures of ≥ 74 °C for 3 min higher than suggested temperatures for scrambled eggs (22% solids) and plain whole egg. Additionally, the ground beef (80:20%) matrix was found to be inhibitory for full toxin activity in the mouse bioassay while retaining some activity in both the cell free translation assay and Vero cell culture cytotoxicity assay.
Topics: Abrin; Animals; Biological Availability; Cell Survival; Chlorocebus aethiops; Eggs; Female; Food Contamination; Food Handling; Mice; Milk; Red Meat; Temperature; Toxins, Biological; Vero Cells
PubMed: 30513721
DOI: 10.3390/toxins10120502 -
Toxins Mar 2019Ricin and abrin are ribosome-inactivating proteins leading to inhibition of protein synthesis and cell death. These toxins are considered some of the most potent and...
Ricin and abrin are ribosome-inactivating proteins leading to inhibition of protein synthesis and cell death. These toxins are considered some of the most potent and lethal toxins against which there is no available antidote. Digital holographic microscopy (DHM) is a time-lapse, label-free, and noninvasive imaging technique that can provide phase information on morphological features of cells. In this study, we employed DHM to evaluate the morphological changes of cell lines during ricin and abrin intoxication. We showed that the effect of these toxins is characterized by a decrease in cell confluence and changes in morphological parameters such as cell area, perimeter, irregularity, and roughness. In addition, changes in optical parameters such as phase-shift, optical thickness, and effective-calculated volume were observed. These effects were completely inhibited by specific neutralizing antibodies. An enhanced intoxication effect was observed for preadherent compared to adherent cells, as was detected in early morphology changes and confirmed by annexin V/propidium iodide (PI) apoptosis assay. Detection of the dynamic changes in cell morphology at initial stages of cell intoxication by DHM emphasizes the highly sensitive and rapid nature of this method, allowing the early detection of active toxins.
Topics: Abrin; Animals; Apoptosis; Cell Survival; Chlorocebus aethiops; HeLa Cells; Humans; Microscopy; Ricin; Vero Cells
PubMed: 30909438
DOI: 10.3390/toxins11030174