Modeling the highly specific ribotoxin recognition of ribosomes

García-Mayoral, Flor; Garcı́a-Ortega, Lucı́a; Álvarez-García, Elisa; Bruix, Marta; Gavilanes, José G.; Pozo, Álvaro Martı́nez del

doi:10.1016/j.febslet.2005.11.027

Cited by 28 publications

(50 citation statements)

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“…However, this cleavage reaction against an isolated SRL-like RNA takes place at rates about 1000-fold slower than when using intact ribosomes [37,46] indicating that additional recognition elements are needed for the optimal ribotoxins' inactivation action against ribosomes. The involvement of the ribosomal context in terms of electrostatic interactions has been suggested not only to justify this recognition and binding enhancement but also to explain the differences found when these toxins are assayed against ribosomes of different origins [18,37]. Electrostatic interactions involving the α-sarcin NH 2 -terminal-β-hairpin have been also suggested to mediate α-sarcin specific recognition of ribosomal proteins [18].…”

Section: Discussionmentioning

confidence: 99%

“…An α-sarcin mutant involving the deletion of this protuberance, α-sarcin Δ (7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22), retained the same conformation as the wildtype protein, as ascertained from its three-dimensional structure in solution [22].…”

Section: Introductionmentioning

confidence: 94%

“…These results were explained by in silico studies that predicted how this NH 2 -terminal β-hairpin could establish essential interactions, mostly of electrostatic nature, with 4 specific ribosomal proteins in order to direct the ribotoxin to the SRL region of the ribosome [18].…”

Section: Introductionmentioning

confidence: 99%

“…This high content of positively charged residues, mostly located at the loops and the NH 2 -terminal β-hairpin, is probably required for recognizing and binding not only to its highly negatively charged target, the SRL rRNA, but also to ribosomal proteins and cellular membranes [18,36,37]. In this context, the role of the positively charged residues located at the NH 2 -terminal β-hairpin (Table 1) in these different but closely related events has been studied in the work herein presented.…”

Section: Introductionmentioning

confidence: 99%

“…Most ribotoxins show a high degree of sequence identity [9][10][11][12] that is also manifested in the three-dimensional structure of the two only ribotoxins studied at this level, restrictocin [13,14] and α-sarcin [15][16][17][18]. Both proteins fold into an α+β structure with a central five-stranded antiparallel β-sheet and an α-helix of almost three turns and display long and unstructured loops ( Fig.1) [ 15,19,20].…”

Section: Introductionmentioning

confidence: 99%

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Role of the basic character of α-sarcin’s NH2-terminal β-hairpin in ribosome recognition and phospholipid interaction

Álvarez-García

Martínez‐del‐Pozo

Gavilanes

2009

Archives of Biochemistry and Biophysics

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Ribotoxins are a family of toxic extracellular fungal RNases that first enter into the cells and then exert a highly specific ribonucleolytic activity on the larger rRNA molecule, leading to protein synthesis inhibition and cell death by apoptosis. α-Sarcin is the best characterized ribotoxin. Previous characterization of a deletion variant of this protein showed that its long NH 2 -terminal β-hairpin is essential for its cytotoxicity. Docking, enzymatic, and lipidprotein interaction studies suggested that this β-hairpin establishes specific interactions with ribosomal proteins and that it is a region involved in the interaction with cell membranes. Consequently, in order to assess the influence of the basic character of this NH 2 -terminal β-hairpin (there are 1 arginine and 4 lysines along its 16 residues) on the ribotoxins cytotoxic ability, five individual mutants substituting these 5 basic residues by glutamic acid were produced, purified to homogeneity, and characterized. Regarding ribosomal recognition, all mutants showed a diminished activity in a cell-free reticulocyte lysate, whereas the activity against an oligoribonucleotide mimicking the sarcin/ricin loop rRNA (SRL) or the homopolymer poly(A) remained unaffected, confirming that the mutated basic residues participate in electrostatic interactions with other ribosomal elements apart from this SRL. The study of the interaction with phospholipid vesicles showed that Lys 17, Arg 22, and, most importantly, Lys 14 and Lys 21, are crucial residues in the first stages of the aggregation phenomenon, where protein-vesicle and protein-protein interactions are required. The data obtained reveal that electrostatic interactions involving basic residues of the β-hairpin are required not only for establishing specific interactions with ribosomal regions other than the SRL but also to explain the ability of the protein to interact with acid phospholipid bilayers.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 94%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Role of the basic character of α-sarcin’s NH2-terminal β-hairpin in ribosome recognition and phospholipid interaction

Álvarez-García

Martínez‐del‐Pozo

Gavilanes

2009

Archives of Biochemistry and Biophysics

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Fungal Ribotoxins

Garcı́a-Ortega

Palacios-Ortega

Martínez‐del‐Pozo

2018

Encyclopedia of Life Sciences

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Fungal ribotoxins constitute a family of extracellular ribonucleases with exquisite specificity against rRNA (ribonucleic acid). They induce apoptotic death of cells after inhibiting protein translation. Ribosomes become functionally incompetent because ribotoxins cleave one single phosphodiester bond, located at a unique and universally conserved loop, needed for elongation factors function. As secreted proteins, ribotoxins need to cross the membrane of their target cells in order to exert their catalytic activity, and they do it without receptor mediation. Using lipid model systems, it has been shown that they are able to enter cells with membranes enriched in acidic phospholipids. Both membrane‐interacting and ribosomal‐recognition activities are characterised by distinct structural features. Even though the natural function of ribotoxins is not known yet, their production by entomopathogenic fungi has suggested their insecticidal role. After decades of detailed study, the biotechnological potential of ribotoxins in pest control and as antitumour agents is becoming evident. Key Concepts Ribotoxins are extremely specific ribonucleases targeted against ribosomes. Ribotoxins are produced by fungi, some of them entomopathogens. They show a high degree of structural conservation, including the local arrangement of the active site residues. Cleavage of a single rRNA phosphodiester bond leads to cell death by inhibiting translation. Ribotoxins are cyclising RNases because they follow a general acid–base mechanism with production of a 2′,3′‐cyclic intermediate. Ribotoxins must first enter their target cells to exert their lethal action. Cell entrance is possible in cells with membranes enriched in acidic phospholipids and altered permeability. Ribotoxins are optimal candidates to be employed as pest control agents and in antitumour immunotoxins.

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RNA toxins: mediators of stress adaptation and pathogen defense

et al. 2011

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RNA toxins are a group of enzymes primarily synthesized by bacteria, fungi, and plants that either cleave or depurinate RNA molecules. These proteins may be divided according to their RNA substrates: ribotoxins are nucleases that cleave ribosomal RNA (rRNA), ribosome inactivating proteins are glycosidases that remove a base from rRNA, messenger RNA (mRNA) interferases are nucleases that cleave mRNAs, and anticodon nucleases cleave transfer RNAs (tRNAs). These modifications to the RNAs may substantially alter gene expression and translation rates. Given that some of these enzymes cause cell death, it has been suggested that they function mainly in defense, either to kill competing cells or to elicit suicide and thereby limit pathogen spread from infected cells. Although good correlations have been drawn between their enzymatic functions and toxicity, recent work has shown that some RNA toxins cause apoptosis in the absence of damage to RNA and that defense against pathogens can be achieved without host cell death. Moreover, a decrease in cellular translation rate, insufficient to cause cell death, allows some organisms to adapt to stress and environmental change. Although ascribing effects observed in vitro to the roles of these toxins in nature has been challenging, recent results have expanded our understanding of their modes of action, and emphasized the importance of these toxins in development, adaptation to stress and defense against pathogens.

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Modeling the highly specific ribotoxin recognition of ribosomes

Cited by 28 publications

References 45 publications

Role of the basic character of α-sarcin’s NH2-terminal β-hairpin in ribosome recognition and phospholipid interaction

Role of the basic character of α-sarcin’s NH2-terminal β-hairpin in ribosome recognition and phospholipid interaction

Fungal Ribotoxins

RNA toxins: mediators of stress adaptation and pathogen defense

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