Heterologous Expression of the Cytotoxin Restrictocin in Aspergillus nidulans and Aspergillus niger

Brandhorst, T. Tristan; Yang, Ruitang; Kenealy, William R.

doi:10.1006/prep.1994.1068

Cited by 17 publications

(19 citation statements)

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“…The phialides of the conidiophores and the emerging, immature conidia of A. restrictus were not eaten during a particular time frame. A dramatic buildup of restrictocin is seen upon the surfaces of these structures during this same time frame (Brandhorst & Kenealy, 1992). This suggests an involvement of the restrictocin in the deterrence of insect feeding.…”

Section: Discussionmentioning

confidence: 65%

“…Cell-associated restrictocin reaches a peak concentration of 18 pg per agar plate culture (Brandhorst & Kenealy, 1992). At this time, there are an average of 1 x 10' conidiophores per plate and approximately 60 phialides per conidiophore.…”

Section: Estimation Of Restrictocin Content Of Differentiated Structuresmentioning

confidence: 99%

“…The mean length and diameter of the phialides, taken from both literature values (Frey et al, 1979) and microscopic measurements by the authors, were used to calculate the approximate volume and weight of the average phialide (assuming a cylindrical shape). The concentration of restrictocin upon the phialides could be estimated given the 10 % relative humidity for the Sit, peamais and 40 quantity of restrictocin present on agar plate cultures of A. restrictus (Brandhorst & Kenealy, 1992).…”

Section: Fungal Culturesmentioning

confidence: 99%

“…The production of restrictocin by A . restrictHs is localized to the conidiophores and phialides during conidium formation (Brandhorst & Kenealy, 1992). It has been suggested that the protein toxin localization could deter insects from feeding until conidia are fully formed, after which time fungus-feeding insects could externally or internally carry spores to new resources for the fungus (Brandhorst & Kenealy, 1992).…”

Section: Introductionmentioning

confidence: 99%

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The ribosome-inactivating protein restrictocin deters insect feeding on Aspergillus restrictus

1996

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The fungus-feeding beetle, Carlpophilus freemani, consumed equal quantities of young mycelia, fewer phialides bearing mature spores and much fewer phialides bearing developing spores of Aspergillus restrictus compared to those of Aspewillus nidulans when tested in diet choice assays. The degree to which specific fungal structures were consumed was inversely related to the localization of high levels of restrictocin, a ribosome-inactivating protein, to those structures. Pure restrictocin added to the insect diet at 1000 p.p.m. killed 385 O/ O of C. fbemani larvae and 62.5 O/ O of SpoCroptera frugiperda larvae in 48 h, but did not affect C. freemsni adults or Helicowerpa zea larvae over the same interval. In diet choice assays, 1000 p.p.m. of restrictocin deterred feeding by adult C. freemani and Sitophilus zeamais compared to control diets. Thus, restrictocin production and localization may have a natural defensive role against insect feeding at times critical to spore formation by A. restrictus, and may have potential as an insect control agent.

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Section: Discussionmentioning

confidence: 65%

Section: Estimation Of Restrictocin Content Of Differentiated Structuresmentioning

confidence: 99%

Section: Fungal Culturesmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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The ribosome-inactivating protein restrictocin deters insect feeding on Aspergillus restrictus

1996

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“…These mutational studies have revealed the involvement of several residues, which are conserved among the different microbial extracellular RNases, in catalysis. Thus, it is well known that His137 and Glu96 are the only residues that are essential for the cleavage reaction performed by α-sarcin [35,45,[70][71][72][73][74][75], whereas His-50, Tyr-48, Arg-121, and Leu-145 mostly contribute to the stabilization of the transition state [45][46][47][48], as stated above. Most of these residues are located in the central β-sheet and their side-chains point towards the concave face of the protein structure where the substrate is supposed to dock [29].…”

Section: Structural Featuresmentioning

confidence: 85%

The Therapeutic Potential of Fungal Ribotoxins

Carreras-Sangrà

Álvarez-García²,

Herrero-Galán³

et al. 2008

CPB

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Ribotoxins constitute a family of toxic extracellular fungal RNases that exert a highly specific activity on a conserved region of the larger molecule of rRNA, known as the sarcin-ricin loop. This cleavage of a single phosphodiester bond inactivates the ribosome and leads to protein synthesis inhibition and cell death. In addition to this ribonucleolytic activity, ribotoxins can cross lipid membranes in the absence of any known protein receptor. This ability is due to their capacity to interact with acid phospholipid-containing membranes. Both activities together explain their cytotoxic character, being rather specific when assayed against some transformed cell lines. The determination of highresolution structures of some ribotoxins, the characterization of a large number of mutants, and the use of lipid model vesicles and transformed cell lines have been the tools used for the study of their mechanism of action at the molecular level. The present knowledge suggests that wild-type ribotoxins or some modified variants might be used in human therapies. Production of hypoallergenic mutants and immunotoxins designed against specific tumors stand out as feasible alternatives to treat some human pathology in the midterm future.3

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Role of histidine-50, glutamic acid-96, and histidine-137 in the ribonucleolytic mechanism of the ribotoxin ?-sarcin

et al. 1999

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alpha-Sarcin is a ribotoxin secreted by the mold Aspergillus giganteus that degrades the ribosomal RNA by acting as a cyclizing ribonuclease. Three residues potentially involved in the mechanism of catalysis--histidine-50, glutamic acid-96, and histidine-137--were changed to glutamine. Three different single mutation variants (H50Q, E96Q, H137Q) as well as a double variant (H50/137Q) and a triple variant (H50/137Q/E96Q) were prepared and isolated to homogeneity. These variants were spectroscopically (circular dichroism, fluorescence emission, and proton nuclear magnetic resonance) characterized. According to these results, the three-dimensional structure of these variants of alpha-sarcin was preserved; only very minor local changes were detected. All the variants were inactive when assayed against either intact ribosomes or poly(A). The effect of pH on the ribonucleolytic activity of alpha-sarcin was evaluated against the ApA dinucleotide. This assay revealed that only the H50Q variant still retained its ability to cleave a phosphodiester bond, but it did so to a lesser extent than did wild-type alpha-sarcin. The results obtained are interpreted in terms of His137 and Glu96 as essential residues for the catalytic activity of alpha-sarcin (His137 as the general acid and Glu96 as the general base) and His50 stabilizing the transition state of the reaction catalyzed by alpha-sarcin.

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Heterologous Expression of the Cytotoxin Restrictocin in Aspergillus nidulans and Aspergillus niger

Cited by 17 publications

References 0 publications

The ribosome-inactivating protein restrictocin deters insect feeding on Aspergillus restrictus

The ribosome-inactivating protein restrictocin deters insect feeding on Aspergillus restrictus

The Therapeutic Potential of Fungal Ribotoxins

Role of histidine-50, glutamic acid-96, and histidine-137 in the ribonucleolytic mechanism of the ribotoxin ?-sarcin

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