Pseudomonas toxin

Fizgerald, D., Morris, R.E., and Saclingcr, C.B. (1980) Receptor-mediated internalization of pseudomonas toxin by mouse fibroblasts. Cell 21, 867. 

Anti-tumour monoclonal antibodies can also be used to deliver toxins to tumour sites. Toxins conjugated to therapeutic antibodies include ricin, pokeweed toxin, Pseudomonas toxin and... 

Some well-known inhibitors of prokaryotic translation include streptomycin, erythromycin, tetracydine, and chloramphenicol. Inhibitors of eukaryotic translation include cycloheximide and Diphtheria and Pseudomonas toxins. 

Like diphtheria toxin. Pseudomonas aeruginosa exotoxin A requires low pH to act (FitzGerald ef al., 1980). In spite of this, it has not been possible to induce translocation of Pseudomonas toxin across the surface membrane by exposure to low pH. It appears that the toxin must be transported beyond the endosomes, possibly to the trans-Golgi network or even to the endoplasmic reticulum to find conditions required for translocation (Chaudhary et al., 1990). In fact domain III ends with an amino acid sequence that (after removal of a terminal... 

Ricin, modeccin, diphtheria toxin, or Pseudomonas toxin induces apoptosis in human lymphoma U937 cells through an increase in caspase-3 and caspase-6 activities, but not caspase-1 activity... 

Komatsu, N., Oda, T. and Muramatsu, T. (1998) Involvement of both caspase-like proteases and serine proteases in apoptotic cell death induced by ricin, modeccin, diphtheria toxin, and pseudomonas toxin. J Biochem (Tokyo), 124, 1038-1044. 

Lee, H. Iglewski, W. J. (1984). Cellular ADP-ribosyltransferase with the same mechanism of action as diphtheria toxin and Pseudomonas toxin A. Proc. Natl. Acad. Sci. USA 81, 2703-2707. 

In a series of experiments designed to characterize the comparative mechanisms of Pseudomonas and Shiga toxin transfer from the Golgi complex to the ER, investigators first sequenced the respective targeting subunits. The C-terminal 24 amino acids of the B subunits of Pseudomonas toxin and Shiga toxin are shown below ... 

C-termlnal 24 amino acids of Pseudomonas toxin B subunit KEQAISALPD YASQPGKPPR KDEL... 

From Inspection of these sequences, what is the probable targeting receptor for transfer of Pseudomonas toxins from the Golgi apparatus to the ER ... 

ADP-ribosylation of eukaryotic elongation factor-2 (EF-2) is recognized as the essential biochemical event in the intoxication of cells by both diphtheria toxin and Pseudomonas toxin A. Both toxins catalyze the transfer of the adenosine 5 -diphosphate ribosyl (ADPR) moiety of nicotinamide adenine dinucleotide (NAD) onto EF-2 [1,2]. The resultant ADPR-EF-2 complex is inactive in protein synthesis. When EF-2 becomes rate limiting, protein synthesis ceases and the intoxicated cell dies. 

NT = not tested Pseudomonas toxin A antisera does not neutralize diphtheria toxin fragment A [10]... 

The Effect of Antisera on the Enzyme Activity of Pseudomonas Toxin A, Fragment A of Diphtheria Toxin, and pyBHK ADP-Ribosyltransferase... 

Fig. 1. Separation of pyBHK EF-2 from the cellular ADP-ribosyltransferase by immunoabsorbant chromatography. A 50 jug sample of an EF-2 preparation containing the cellular transferase activity was applied to a 250 jul column of Sepharose 4B resin coupled to Pseudomonas toxin A antibody. Unbound sample was washed from the resin with phosphate-buffered saline containing 1 vaM DTT (PBS). Protein bound to the immunoabsorbant was dissociated and eluted in 1 M propionic add containing 1 mAf DTT. Acid-containing fractions were immediately neutralized with 2 M Tris-Base. A portion of each fraction was assayed for EF-2 ( — ) by the transfer of [ C]-adenosine from NAD to the EF-2 in the presence of diphtheria toxin fragment A [6]. A second portion of each fraction was assayed for pyBHK ADP-ribosyltransferase activity (o—o) by the transfer of [ C]-adenosine from NAD to EF-2 in our standard cellular transferase reaction mixture [7] which was supplemented with 4 jug of a pyBHK EF-2 preparation which lacked its own endogenous transferase activity. Acid precipitable radioactivity was detected using a liquid scintillation counter...

Fig. 2. Western blot analysis of protein bound to immunoabsorbant columns. PyBHK EF-2 preparations containing cellular ADP-ribosyltransferase activity were chromatographed on Sepharose 4B coupled to Pseudomonas toxin A antibody or pyBHK ADP-ribosyltransferase antibody. After washing unbound material from the resin with PBS, the bound protein was eluted from the columns with 1 M propionic acid, concentrated under vacuum and subjected to electrophoresis on a 7.5% SDS-polyacrylamide slab gel. In addition. Pseudomonas toxin A standards were subjected to electrophoresis on the gel. Following electroblotting of the proteins from the ge) to a nitrocellulose sheet, the sheet was sectioned and reacted with Pseudomonas toxin A antibody (A-Q or with pyBHK ADP-ribosyltransferase antibody iP-F). Pseudomonas toxin A (A) protein from Pseudomonas toxin A antibody coupled immunoabsorbant B) protein from pyBHK ADP-ribosyltransferase antibody coupled immunoabsorbant (C, D) protein from Pseudomonas toxin A antibody-coupled immunoabsorbant (i ) Pseudomonas toxin A F). The numbers represent Mj. X 10" of the mol.wt. standards...

Photolabeling of a limited array of other proteins with [carbonyl- " C]NAD was tested under identical conditions. Table 1 shows that besides fragment A, the ApUp-free form of DT, CRM-45, and activated Pseudomonas toxin incorporated label to much greater extents than did other proteins tested. The amounts of label incorporated by intact DT and by activated Pseudomonas toxin were virtually identical, suggesting similarities in the nicotinamide subsites of these two toxins. 

The mono(ADP-ribosyl)ation, the transfer of the ADP-ribose moiety of NAD to a macromolecule, was discovered by Hayaishi et al. in 1968 as the mechanism of the cytotoxic effect of diphtheria toxin [1], The substrate of this toxin-catalyzed ADP-ribosylation is elongation factor-2. The same reaction is catal)Azed by Pseudomonas toxin. The second bacterial toxin involved in mono(ADP-ribosyl)ation of mammalian cell proteins is cholera toxin, the substrate of which was identified as the guanine nucleotide-binding regulatory component of membrane adenylate cyclase in 1978 [2].E. coli heat-labile enterotoxin is similar to cholera toxin in many respects. 

Bialaphos and tabtoxinine-j8-lactam, a Pseudomonas toxin, both cause the accumulation of high intracellular ammonia concentrations in plant tissues.There is also a substantial body of evidence that this aspect of GS inhibition is important in the herbicidal activity of ppj. s-iQs... 

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