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Cellular and Molecular Life Sciences :... Aug 2006Ribosome-inactivating proteins (RIPs), mostly from plants, are enzymes which depurinate rRNA, thus inhibiting protein synthesis. They also depurinate other... (Review)
Review
Ribosome-inactivating proteins (RIPs), mostly from plants, are enzymes which depurinate rRNA, thus inhibiting protein synthesis. They also depurinate other polynucleotide substrates. The biological activity of RIPs is not completely clarified, and sometimes independent of the inhibition of protein synthesis. There are differences in the cytotoxicity of RIPs and, consequently, in their toxicity to animals. Some RIPs are potent toxins, the best known being ricin, a potential biological weapon. New toxins have recently been identified. RIPs cause apoptotic and necrotic lesions, and induce production of cytokines causing inflammation. RIPs are potentially useful in agriculture and medicine because (i) they have antiviral activity and (ii) they are used for the preparation of conjugates with antibodies ('immunotoxins') or other carriers, rendering them specifically toxic to the cell target of the carrier, which may be helpful in therapy. The distribution, mechanism of action and role in nature of RIPs are not completely understood, and we can expect several future developments in their practical application.
Topics: Animals; Antineoplastic Agents; Humans; N-Glycosyl Hydrolases; Neurons; Plant Proteins; RNA, Plant; RNA, Ribosomal; RNA, Ribosomal, 28S; Ribosomes
PubMed: 16799768
DOI: 10.1007/s00018-006-6078-7 -
Virologica Sinica Dec 2011Many plants contain ribosome inactivating proteins (RIPs) with N-glycosidase activity, which depurinate large ribosomal RNA and arrest protein synthesis. RIPs so far... (Review)
Review
Many plants contain ribosome inactivating proteins (RIPs) with N-glycosidase activity, which depurinate large ribosomal RNA and arrest protein synthesis. RIPs so far tested inhibit replication of mRNA as well as DNA viruses and these proteins, isolated from plants, are found to be effective against a broad range of viruses such as human immunodeficiency virus (HIV), hepatitis B virus (HBV) and herpes simplex virus (HSV). Most of the research work related to RIPs has been focused on antiviral activity against HIV; however, the exact mechanism of antiviral activity is still not clear. The mechanism of antiviral activity was thought to follow inactivation of the host cell ribosome, leading to inhibition of viral protein translation and host cell death. Enzymatic activity of RIPs is not limited to depurination of the large rRNA, in addition they can depurinate viral DNA as well as RNA. Recently, Phase I/II clinical trials have demonstrated the potential use of RIPs for treating patients with HIV disease. The aim of this review is to focus on various RIPs from plants associated with anti-HIV activity.
Topics: Animals; Antiviral Agents; Down-Regulation; HIV; Humans; Plant Proteins; Plants; Ribosome Inactivating Proteins; Virus Replication
PubMed: 22160935
DOI: 10.1007/s12250-011-3223-8 -
Toxicology Letters Sep 2016The plant-derived toxins ricin and abrin, operate by site-specific depurination of ribosomes, which in turn leads to protein synthesis arrest. The clinical manifestation... (Comparative Study)
Comparative Study
The plant-derived toxins ricin and abrin, operate by site-specific depurination of ribosomes, which in turn leads to protein synthesis arrest. The clinical manifestation following pulmonary exposure to these toxins is that of a severe lung inflammation and respiratory insufficiency. Deciphering the pathways mediating between the catalytic activity and the developing lung inflammation, requires a quantitative appreciation of the catalytic activity of the toxins, in-vivo. In the present study, we monitored truncated cDNA molecules which are formed by reverse transcription when a depurinated 28S rRNA serves as template. We found that maximal depurination after intranasal exposure of mice to 2LD50 ricin was reached 48h, where nearly 40% of the ribosomes have been depurinated and that depurination can be halted by post-exposure administration of anti-ricin antibodies. We next demonstrated that the effect of ricin intoxication on different cell types populating the lungs differs greatly, and that outstandingly high levels of damage (80% depurination), were observed in particular for pulmonary epithelial cells. Finally, we found that the magnitude of depurination induced by the related plant-derived toxin abrin, was significantly lower in comparison to ricin, and can be attributed mostly to reduced depurination of pulmonary epithelial cells by abrin. This study provides for the first time vital information regarding the scope and timing of the catalytic performance of ricin and abrin in the lungs of intact animals.
Topics: Abrin; Abrus; Administration, Intranasal; Animals; Antitoxins; Cytotoxins; DNA, Complementary; Female; Flow Cytometry; Lethal Dose 50; Lung; Mice; Pneumonia; Poisoning; Protein Synthesis Inhibitors; Purines; RNA, Ribosomal, 28S; Respiratory Insufficiency; Respiratory Mucosa; Ribosomes; Ricin; Ricinus
PubMed: 27298272
DOI: 10.1016/j.toxlet.2016.06.003 -
Toxins Apr 2018Ribosome inactivating proteins (RIPs) form a vast family of hundreds of toxins from plants, fungi, algae, and bacteria. RIP activities have also been detected in animal...
Ribosome inactivating proteins (RIPs) form a vast family of hundreds of toxins from plants, fungi, algae, and bacteria. RIP activities have also been detected in animal tissues. They exert an N-glycosydase catalytic activity that is targeted to a single adenine of a ribosomal RNA, thereby blocking protein synthesis and leading intoxicated cells to apoptosis. In many cases, they have additional depurinating activities that act against other nucleic acids, such as viral RNA and DNA, or genomic DNA. Although their role remains only partially understood, their functions may be related to plant defense against predators and viruses, plant senescence, or bacterial pathogenesis.
Topics: Animals; Humans; Plant Physiological Phenomena; Ribosome Inactivating Proteins; Toxins, Biological
PubMed: 29669991
DOI: 10.3390/toxins10040160 -
Current Topics in Microbiology and... 2012Ricin and Shiga toxins designated as ribosome inactivating proteins (RIPs) are RNA N-glycosidases that depurinate a specific adenine (A₄₃₂₄ in rat 28S rRNA) in... (Review)
Review
Ricin and Shiga toxins designated as ribosome inactivating proteins (RIPs) are RNA N-glycosidases that depurinate a specific adenine (A₄₃₂₄ in rat 28S rRNA) in the conserved α-sarcin/ricin loop of the large rRNA, inhibiting protein synthesis. Evidence obtained from a number of studies suggests that interaction with ribosomal proteins plays an important role in the catalytic activity and ribosome specificity of RIPs. This review summarizes the recent developments in identification of the ribosomal proteins that interact with ricin and Shiga toxins and the principles governing these interactions.
Topics: Enzyme Activation; Protein Binding; Ribosomal Proteins; Ribosome Inactivating Proteins; Ribosomes; Ricin; Shiga Toxins
PubMed: 21910078
DOI: 10.1007/82_2011_174 -
Phytochemistry Oct 2022Ribosome inactivating proteins (RIPs) are rRNA N-glycosylases (EC 3.2.2.22) best known for hydrolyzing an adenine base from the conserved sarcin/ricin loop of ribosomal... (Review)
Review
Ribosome inactivating proteins (RIPs) are rRNA N-glycosylases (EC 3.2.2.22) best known for hydrolyzing an adenine base from the conserved sarcin/ricin loop of ribosomal RNA. Protein translation is inhibited by ribosome depurination; therefore, RIPs are generally considered toxic to cells. The expression of some RIPs is upregulated by biotic and abiotic stress, though the connection between RNA depurination and defense response is not well understood. Despite their prevalence in approximately one-third of flowering plant orders, our knowledge of RIPs stems primarily from biochemical analyses of individuals or genomics-scale analyses of small datasets from a limited number of species. Here, we performed an unbiased search for proteins with RIP domains and identified several-fold more RIPs than previously known - more than 800 from 120 species, many with novel associated domains and physicochemical characteristics. Based on protein domain configuration, we established 15 distinct groups, suggesting diverse functionality. Surprisingly, most of these RIPs lacked a signal peptide, indicating they may be localized to the nucleocytoplasm of cells, raising questions regarding their toxicity against conspecific ribosomes. Our phylogenetic analysis significantly extends previous models for RIP evolution in plants, predicting an original single-domain RIP that later evolved to acquire a signal peptide and different protein domains. We show that RIPs are distributed throughout 21 plant orders with many species maintaining genes for more than one RIP group. Our analyses provide the foundation for further characterization of these new RIP types, to understand how these enzymes function in plants.
Topics: Phylogeny; Plant Proteins; Protein Sorting Signals; RNA, Ribosomal; Ribosome Inactivating Proteins; Ribosomes
PubMed: 35934106
DOI: 10.1016/j.phytochem.2022.113337 -
Chromosoma Apr 1979C-banding of chromosomes involves the differential solubilization of fragmented DNA from euchromatin by three sequential treatments: 1. Acid, 2. Mild base, 3. Hot salt....
C-banding of chromosomes involves the differential solubilization of fragmented DNA from euchromatin by three sequential treatments: 1. Acid, 2. Mild base, 3. Hot salt. The data indicate solubilization is effected by 1) depurination, 2) DNA denaturation, 3) chain breakage of the depurinated sites respectively in the three treatments. Conditions were found wherein each treatment in proper sequence was necessary for C-banding and the appropriate chemical reactions were measured in these treatment conditions. The acid treatment (0.2 N HCl) depurinates chromosomal DNA at the rate of 0.26 x 10(-6) purines/dalton min to an alkaline molecular weight of 10(5) daltons but does not break the depurinated sites. Bleomycin can substitute for acid as a base removing agent. Sodium borohydride, by reducing the depurinated sugar's aldehyde thereby preventing chain breakage by the beta-elimination reaction, reversibly inhibits DNA-extraction. Chain breakage at the DNA's apurinic sites occurs not in the 2 min mild alkali treatment where the half-life for breakage is 26 min but in the 18 h hot salt treatment where the half-life for chain breakage is 1-2 h. Most of the DNA extraction occurs in the hot salt as 10(5) dalton fragments as measured in formamide gradients. Bleomycin is introduced as a substitute for HCl; it removes nitrogenous bases from DNA in situ while better preserving the morphology of the final C-banded chromosomes.
Topics: Animals; Azure Stains; Bleomycin; Cell Line; Chromosome Banding; DNA; Hot Temperature; Mice; Molecular Weight; Purines
PubMed: 88314
DOI: 10.1007/BF00293235 -
Nucleic Acids Research Aug 1999DNA fragments that show a pattern of differential expression on differential display gels must be eluted from the gel matrix and reamplified to enable further analysis....
DNA fragments that show a pattern of differential expression on differential display gels must be eluted from the gel matrix and reamplified to enable further analysis. Elution is usually achieved by heating excised gel slices in a small volume of either water or TE. Here we show that this elution step can adversely affect the ability of the eluted DNA to act as a template for PCR reamplification, probably via the process of depurination. Simply switching to an elution solvent designed to minimise depurination (PCR buffer) facilitates the elution of intact DNA fragments. This improvement is likely to be most beneficial when eluting higher molecular weight fragments (e.g. those >500 bp), in situations where the amount of DNA in an excised gel slice is limited or when contaminating differential display products co-migrate with the differentially expressed product.
Topics: DNA; Diffusion; Electrophoresis, Polyacrylamide Gel; Kinetics; Polymerase Chain Reaction; Purines; Temperature; Templates, Genetic
PubMed: 10454625
DOI: 10.1093/nar/27.15.e6 -
Analytical Chemistry Apr 2009Ricin is a family member of the lethal ribosome-inactivating proteins (RIP) found in plants. Ricin toxin A-chain (RTA) from castor beans catalyzes the hydrolytic...
Ricin is a family member of the lethal ribosome-inactivating proteins (RIP) found in plants. Ricin toxin A-chain (RTA) from castor beans catalyzes the hydrolytic depurination of a single base from a GAGA tetraloop of eukaryotic rRNA to release a single adenine from the sarcin-ricin loop (SRL). Protein synthesis is inhibited by loss of the elongation factor binding site resulting in cell death. We report a sensitive coupled assay for the measurement of adenine released from ribosomes or small stem-loop RNAs by RTA catalysis. Adenine phosphoribosyl transferase (APRTase) and pyruvate orthophosphate dikinase (PPDK) convert adenine to ATP for quantitation by firefly luciferase. The resulting AMP is cycled to ATP to give sustained luminescence proportional to adenine concentration. Subpicomole adenine quantitation permits the action of RTA on eukaryotic ribosomes to be followed in continuous, high-throughput assays. Facile analysis of RIP catalytic activity will have applications in plant toxin detection, inhibitor screens, mechanistic analysis of depurinating agents on oligonucleotides and intact ribosomes, and in cancer immunochemotherapy. Kinetic analysis of the catalytic action of RTA on rabbit reticulocyte 80S ribosomes establishes a catalytic efficiency of 2.6 x 10(8) M(-1) s(-1), a diffusion limited reaction indicating catalytic perfection even with large reactants.
Topics: Adenine; Adenosine Monophosphate; Animals; Inverted Repeat Sequences; Kinetics; Luciferases, Firefly; Luminescent Measurements; Nucleic Acid Hybridization; Purines; Pyruvate, Orthophosphate Dikinase; RNA; Reticulocytes; Ribosome Subunits, Large, Eukaryotic; Ribosomes; Ricin; Sensitivity and Specificity; Time Factors
PubMed: 19364139
DOI: 10.1021/ac8026433 -
Virulence Nov 2013Ribosome-inactivating proteins (RIPs) were first isolated over a century ago and have been shown to be catalytic toxins that irreversibly inactivate protein synthesis.... (Review)
Review
Ribosome-inactivating proteins (RIPs) were first isolated over a century ago and have been shown to be catalytic toxins that irreversibly inactivate protein synthesis. Elucidation of atomic structures and molecular mechanism has revealed these proteins to be a diverse group subdivided into two classes. RIPs have been shown to exhibit RNA N-glycosidase activity and depurinate the 28S rRNA of the eukaryotic 60S ribosomal subunit. In this review, we compare archetypal RIP family members with other potent toxins that abolish protein synthesis: the fungal ribotoxins which directly cleave the 28S rRNA and the newly discovered Burkholderia lethal factor 1 (BLF1). BLF1 presents additional challenges to the current classification system since, like the ribotoxins, it does not possess RNA N-glycosidase activity but does irreversibly inactivate ribosomes. We further discuss whether the RIP classification should be broadened to include toxins achieving irreversible ribosome inactivation with similar turnovers to RIPs, but through different enzymatic mechanisms.
Topics: Bacterial Toxins; Humans; Poisons; Protein Synthesis Inhibitors; RNA, Ribosomal, 28S; Ribosome Inactivating Proteins
PubMed: 24071927
DOI: 10.4161/viru.26399