Toxin structure

We started feeding experiments using C-labeled acetate long before the establishment of the toxin structures with a hope that the analysis of the C-NMR... 

In addition to variations in toxin structure, the nature of the binding site and the medium in which binding occurs both influence the observed behavior of the toxins and can be systematically varied to study the toxin/binding site interaction. These factors will not be discussed at length here, but should be remembered as complications in comparing different data sets. 

The PSP toxins represent a real challenge to the analytical chemist interested in developing a method for their detection. There are a great variety of closely related toxin structures (Figure 1) and the need exists to determine the level of each individually. They are totally non-volatile and lack any useful UV absorption. These characteristics coupled with the very low levels found in most samples (sub-ppm) eliminates most traditional chromatographic techniques such as GC and HPLC with UVA S detection. However, by the conversion of the toxins to fluorescent derivatives (J), the problem of detection of the toxins is solved. It has been found that the fluorescent technique is highly sensitive and specific for PSP toxins and many of the current analytical methods for the toxins utilize fluorescent detection. With the toxin detection problem solved, the development of a useful HPLC method was possible and somewhat straightforward. 

One of the first applications of the HPLC method was the investigation of differences in toxin profiles between shellfish species from various localities ( ). It became apparent immediately that there were vast differences in these toxin profiles even among shellfish from the same beach. There were subtle differences between the various shellfish species, and butter clams had a completely different suite of toxins than the other clams and mussels. It was presumed that all of the shellfish fed on the same dinoflagellate population, so there must have been other factors influencing toxin profiles such as differences in toxin uptake, release, or metabolism. These presumptions were strengthened when toxin profiles in the littleneck clam (Prototheca Staminea) were examined. It was found that, in this species, none of the toxin peaks in the HPLC chromatogram had retention times that matched the normal PSP toxins. It was evident that some alteration in toxin structure had occurred that was unique in this particular shellfish species. 

Work with European toxic cyanobacteria was partially supported by a NATO collaborative research grant between W.W. Carmichael and G.A. Codd, University of Dundee, Scotland, and O.M. Skulberg, Norwegian Water Research Institute, Oslo, Norway. Toxin structure work on European and North American peptide toxins is supported in part by U.S. AMRDC contract DAMD17-87-C-7019 to W.W. Carmichael. Portions of the work represent part of the Ph.D. dissertation research of N.A. Mahmood and E.G. Hyde. Their work was supported in part by fellowship support from the Biomedical Ph.D. Program, Wright State University. 

More recent studies on the folded toxin structure by Norton and colleagues have utilized h- and C-NMR techniques (19,20). By using 2D-FT-NMR, it was possible to localize a four stranded, antiparallel )5-pleated sheet "backbone structure in As II, Ax I, and Sh I (21,22), In addition, Wemmer et al. (23) have observed an identical )5-pleated structure in Hp II. No a-helix was observed in these four variants. In the near future, calculated solution conformations of these toxins, utilizing distance measurements from extracted Nuclear Overhauser Enhancement (NOE) effects should greatly stimulate structure-activity investigations. 

Disulfide bonds, howpver, are important in maintaining the particular toxin structure and have been shown to be essential for toxicity. When all four disulfide bonds are reduced and alkylated, the neurotoxin loses its toxicity 4). 

Kem WR. Sea anemone toxins structure and action. In Hessinger D, Lenhoff H, eds. The Biology of Nematocysts. New York Academic Press, 1988 375-405. 

Di Pierro M, Lu R, Uzzau S, Wang W, Margaretten K, Pazzani C, Maimone F, Fasano A (2001) Zonula occludens toxin structure-function analysis. J Biol Vhem 276 19160-19165 Doyle LA, Yang W, Abruzzo LY, Krogmann T, Gao Y, Rishi AK, Ross DD (1998) A multidrug resistance transporter from human MCF-7 breast cancer cells. Proc Natl Acad Sci USA 95 15665-15670... 

Many animal peptide toxins have been characterised by NMR spectroscopy, including examples from species as diverse as marine sponges to mammals. Although these peptides vary significantly in their sequence, structure and function, they do share common features, such as the structural motifs present and their activities. For example, many toxins target ion channels or have antimicrobial activity. In this section, we have chosen examples from the vast array of available peptide toxin structures to illustrate this structural and functional diversity and similarity. 

The compound made by this reaction has almost, but not exactly, the spider toxin structure. The extra groups in brown are protecting groups, and prevent unwanted side-reactions at the other amine and phenol functional groups. We will discuss protecting groups in detail in Chapters 24 and 25. 

Td and each fraction has 2 chemical components. Collectively these are the AAL-toxins. Structurally in T j, Rj and R3 -... 

Like some other membrane active proteins and peptides [95-97], Cry toxins bind to the cell siuface in a water soluble form, followed by an irreversible conformational change converting into a form capable of inserting into the membrane [35]. The Cry toxin structures revealed that putative membrane-spaiming amphipathic helices located in domain I might be involved in pore formation [30,31]. Since the amphipathic helices predicted to span the membrane are buried in the helical bundle in domain I, a conformational change was predicted to expose a relatively non-polar/hydrophobic hairpin composed of helices a4 and a5 to initiate membrane insertion [30,31]. 

The application of RRFs has not been explored for the LC-FL methods for saxitoxins using pre-or postcolumn oxidation. The yield of oxidation product and FL RF are very dependent on the toxin structure and the details of the instrument setup. Therefore, use of RRFs may not be feasible. This is unfortunate because it is a huge task to prepare and maintain stocks of CRMs for the wide range of saxitoxin congeners that can be found in seafoods. ... 

C-7 hydroxyl (7-0-acylation). The C-1 esters are formed within algae and the 7-0-acyl esters are formed within shellfish. There are many reports of not fully characterized variations to this general scheme and for most purposes, it can be accepted that when an OA diol ester has been reported, a DTX-1 and DTX-2 ester might also exist. For the purpose of simplicity, many, but not all, the elucidated OA group toxin structures are shown in Figure 10.2. 

Among these different biopesticides, bacterial biopesticides are the most intensively studied and widely used. Several insect pathogenic bacteria are known to produce proteins toxic to certain insects. Bacillus thurin ensis (Bt) is the most well-known bacterium for its potent insecticidal proteins. These proteins are highly specific to certain orders of insects. Insects sensitive to Bt include those of Coleoptera, Diptera, and Lepidoptera. Bacillus sphaericus and Clostridium bifermentans are known for their mosquitocidal proteins. Paenibadllus popilliae produces a scarab active toxin structurally similar to common insecticidal proteins... 

Robertus J. Toxin structure. In Frankel A, ed. Immunotoxins. Boston, Mass Kluwer Academic Publishers 1988 11-24. 

Anthrax Toxin Structure and Fimction NIDR/Leppla... 

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