Toxicity Botulinum neurotoxin

Hanna PA, Jankovic J, Vincent A (1999) Comparison of mouse bioassay and immunoprecipitation assay for botulinum toxin antibodies. J Neurol Neurosurg Psychiatry 66 612-16 Hanson MA, Stevens RC (2000) Cocrystal structure of synaptobrevin-II bound to botulinum neurotoxin type B at 2.0 A resolution. Nat Struct Biol 7 687-92 Harlow ML, Ress D, Stoschek A, Marshall RM, McMahan UJ (2001) The architecture of active zone material at the frog s neuromuscular junction. Nature 409 479-84 Harris JB (1997) Toxic phospholipases in snake venom an introductory review. Symp. zool. Soc. Lond. 70 235-50... 

Lacy DB, Tepp W, Cohen AC, DasGupta BR, Stevens RC (1998) Crystal structure of botulinum neurotoxin type a and implications for toxicity. Nat Struct Biol 5 898-902... 

Clostridium bofulinum type C was firstly isolated by Bengston in 1922 (Bengston, 1922). In 1935, Mason and Robinson reported that C. bofulinum type C produced three different toxic factors, Cl, C2 and D (Mason and Robinson, 1935), although it had been considered at that time that other types of C. bofulinum, types A and B, produced only one antigenic type of the toxin. This was the first use of the term C2 toxin in the literature. Later on, Jansen applied this notion to the toxins produced by C. bofulinum C and Cp strains, which had been classified by immunological cross-neutralization produces Cl, C2 and D toxins and Cp only C2 toxin (Jansen, 1971). Thus, C2 toxin had been thought of as a botulinum neurotoxin until it was purified and characterized in 1980 (Ohishi etal., 1980). 

Prepare a culture of a C2 toxin-producing strain of C botulinum it is preferable to use strains, that do not produce botulinum neurotoxin, because the neurotoxin interferes the determination of toxicity of C2 toxin. Otherwise, use anti-neurotoxin serum to neutralize the neurotoxic activity. 

Although the toxins are produced in large amounts by the relevant bacterial strains and are not difficult to purify, until recently most of them have been available only to a limited number of laboratories. The main reason for this is their extreme toxicity, particularly in the case of the botulinum neurotoxins. 

The seven serotypes of botulinum toxin produced by Clostridium botulinum are the most toxic substances known. They are associated with lethal food poisoning after the consumption of canned foods. This family of toxins was evaluated by the United States as a potential biological weapon in the 1960s and is believed to be an agent that could be used against our troops. Unlike other threat toxins, botulinum neurotoxin appears to cause the same disease after inhalation, oral ingestion, or injection. Death results from skeletal muscle paralysis and resultant ventilatory failure. Because of its extreme toxicity, the toxin typically cannot be identified in body fluids, other than nasal... 

Fig. 4. Organization and fragmentation of botulinum neurotoxin. Botulinum neurotoxin is a protein with a molecular weight of 150,000 to 160,000. It is synthesized as a single chain polypeptide in the Clostridium bacteria. It is nicked by endogenous protease to yield a dichain molecule linked by a disulfide bond. This nicking is usually associated with activation of its toxicity. The nicked molecule is separated into heavy and light chains by reduction of the disulfide bond.

Ilkhchoui Yl, Ghaly RF, Knezevic NN, Candido KD. Central nervous system toxicity after botulinum neurotoxin irqection. Anesth Pain Med... 

Botulism, the deadly food poisoning disease is caused by the growth of various strains of Clostridium botulinum in food. The organism produces a large polypeptide (neurotoxin) which is the most toxic protein known to the human kind. Seven serotypes of botulinum neurotoxins produced by different strains of C. botulinum have been characterized, and serotypes A, B and E are known to cause botulism in humans. Ingestion of food contaminated with the neurotoxin causes flaccid muscle paralysis that can result in patients death. Wound botulism has also been reported where the organism can grow in the wounds, and produces the neurotoxin that causes paralysis. 

Figure 2. Schematic diagram of botulinum neurotoxin showing its light and heavy chains. The two different domains of the heavy chain shaded with different patterns indicate the N-terminal and C-terminal halves (about 50 kDa each). These two domains are believed to play different functional roles during the intoxication process. The light chain has been shown to contain the toxic site.

Treatment of the neurotoxin with metal chelators such as ethylenediamine tetraacetic acid (EDTA) inactivate most (80%) of the enzymatic activity, and large part (80%) of the lost activity can be restored with the addition of Zn " ions activity (Foran et al., 1994). Similar loss of toxic activity of type A botulinum neurotoxin was recently observed in our studies with the neurotransmitter release studies in PC 12 cells (F. -N. Fu, R. Lomneth and B. R. Singh, unpublished data). However, the loss of activity was not restored with the replenishment with Zn " ions. The loss of activity at least in part seems to be caused by the change in the polypeptide folding (Fu and Singh, 1995). 

Botulinum neurotoxins, produced by the anaerobic bacterium Clostridium bo-tulinum, are the most toxic poisons known to man. The neurotoxins are food poisons. Once ingested, the neurotoxin is absorbed through the intestinal mucosal layer into the blood stream. It acts at the neuromuscular junction to inhibit the release of acetylcholine (a neurotransmitter) from nerve endings (Simpson, 1989). The result is the dreaded botulism disease, which is manifested by flaccid muscle paralysis. 

Until recently, there have been only two primary techniques available for the detection of botulinum neurotoxins. The first of these, which is the most widely accepted and sensitive technique for the detection of botulinum neurotoxins in semm and food extracts, is the mouse bioassay (Sakaguchi, 1983). Although the mouse bioassay is the most sensitive method, with the ability to detect less than 5 mouse 50% lethal doses (MLD5os)/mL, the assay takes up to four days to complete and requires a large number of mice if the toxin is to be quantified. In addition, the mouse toxicity results are not in themselves specific specificity is imparted only by carrying out parallel toxin neutralization tests with homologous antisera (Shone et al., 1985). Furthermore, as future modifications are made to botulinum neurotoxins relative to their use as therapeutic agents, quantification relative to its toxicity to mice may not be possible. Thus, despite the apparent sensitivity offered by the mouse bioassay, its use as a routine detection technique for botulinum neurotoxins is not only impractical, but also may be obsolete in some areas of research. 

Classical bacterial exotoxins, such as diphtheria toxin, cholera toxin, clostridial neurotoxins, and the anthrax toxins are enzymes that modify their substrates within the cytosol of mammalian cells. To reach the cytosol, these toxins must first bind to different cell-surface receptors and become subsequently internalized by the cells. To this end, many bacterial exotoxins contain two functionally different domains. The binding (B-) domain binds to a cellular receptor and mediates uptake of the enzymatically active (A-) domain into the cytosol, where the A-domain modifies its specific substrate (see Figure 1). Thus, three important properties characterize the mode of action for any AB-type toxin selectivity, specificity, and potency. Because of their selectivity toward certain cell types and their specificity for cellular substrate molecules, most of the individual exotoxins are associated with a distinct disease. Because of their enzymatic nature, placement of very few A-domain molecules in the cytosol will normally cause a cytopathic effect. Therefore, bacterial AB-type exotoxins which include the potent neurotoxins from Clostridium tetani and C. botulinum are the most toxic substances known today. However, the individual AB-type toxins can greatly vary in terms of subunit composition and enzyme activity (see Table 2). 

M toxin (12S) consists of neurotoxin and non-toxic non-hemaglutynin (NTNH) components. It is produced by all types of C. botulinum, except C. botulinum type G. 

Botulinum toxin is both a medication and a neurotoxin, produced by the bacterium Clostridium botulinum. It is the most toxic protein known. It can be used to treat muscle spasms, and is sold commercially under various names (Botox, Dysport, Myobloc, etc.). Botox Cosmetic and Vistabel are available for cosmetic treatment. The toxin protein consists... 

Although inhalational botulinum intoxication was investigated in other animal species, these studies have not provided specific data on toxin absorption. The behavior of BoNTs in the respiratory tract was only recently investigated. Park and Simpson (2003) studied the properties of pure BoNT/A neurotoxin both in vivo and in vitro using mice and pulmonary cell culture models, respectively. Mean survival times were compared in mice receiving various doses of pure BoNT/A either IN or IP. Pure BoNT/A was found to be a potent intranasal poison, although the toxicity (as determined by mean survival time) associated with IP administration was somewhat higher. Mean survival times in mice were less than 100 (IP) or 600 min (IN) after administration of 0.1 pg pure toxin 75 (IP) or 400 min (IN) for 1 pg toxin and 120 min (IN) for 10 pg toxin (Park and... 

A-B Toxins are bacterial toxins composed of two peptide chains one (B) that binds to the invaded cell surface, and the other (A) containing the toxin which is then taken-up into the cell. Some examples of exotoxins secreted by the bacteria into the surrounding medium and highly toxic to certain tissues are pathogens causing botuiism (Clostridium botulinum), tetanus (Clostridium tetani) and diptheria (Corynebacterium diphtheria. An example of an A-B endotoxin is Vibrio cholerae. Botulinum toxin and tetanus toxin have their main toxic actions on neuronal tissues, so are described at NEUROTOXINS. 

Several toxic proteins act as neurotoxins by disrupting the activity of synapses. (A synapse is a junction between two neurons or between a neuron and a muscle cell.) The pain, tremors, and irritability that result from black widow spider bites are caused by a-latrotoxin (125,000 D). This molecule, a single polypeptide, stimulates a massive release of the neurotransmitter acetylcholine (ACh). In contrast, ACh release is inhibited by botulinum toxin, a mixture of several proteins produced by the bacterium Clostridium botulinum. Botulism, a malady most commonly caused by eating contaminated canned food, is characterized by vomiting, dizziness, and sometimes paralysis and death. A related species,... 

---