Alpha-amanitin

Alpha-amanitin or α-amanitin is a cyclic nonribosomal peptide of eight amino acids. It is possibly the most deadly of all the amatoxins, toxins found in several members of the Amanita genus of mushrooms, one being the Death cap (Amanita phalloides) as well as the Destroying angel, a complex of similar species, principally A. virosa and A. bisporiga. It is also found in the mushrooms Galerina autumnalis and Conocybe filaris. The oral of amanitin is approximately 0.1 mg/kg.

The structure of the polypeptide is atypical of most polypeptides, due to the branching of the amino acid chain. Two modified amino acids (dihydroxylated tryptophan and a sulfinated cysteine) allow the formation of a second "inner loop", as seen in the diagram at right. The "outer loop" is formed by the normal peptide bond of the carboxyl terminus to the amino terminus of the peptide chain.

Scientific use
alpha-Amanitin is an inhibitor of RNA polymerase II. This mechanism makes it a deadly toxin.

alpha-Amanitin can also be used to determine which types of RNA polymerase are present. This is done by testing the sensitivity of the polymerase in the presence of alpha-amanitin. RNA polymerase I is insensitive, RNA pol II is highly sensitive, and RNA pol III is slightly sensitive.

Symptoms of poisoning
Alpha-amanitin has an unusually strong and specific attraction to the enzyme RNA polymerase II. Upon ingestion, it binds to the RNA polymerase II enzyme, effectively causing cytolysis of hepatocytes (liver cells). Few effects are reported within 10 hours; it is not unusual for significant effects to take as much as 24 hours after ingestion, this delay in symptoms making it even more difficult to diagnose as well as dangerous. By then, it is far past the time in which stomach pumping would yield an efficient result. Diarrhea and cramps are the first symptoms, but those pass, giving a false sign of remission. Typically, on the 4th to 5th day, the toxin starts to have severe effects on the liver and kidneys, leading to total system failure in both. Death usually takes place around a week from ingestion.

Around 15% of those poisoned will die in around 10 days progressing through a comatose stage to renal failure, liver failure, hepatic coma, respiratory failure and death. Those who recover are at risk of permanent liver damage. Diagnosis is difficult, centered around the described clinical symptoms as well as alpha-amanitin content excreted in the urine. Urine screening is generally most useful within 48 hours of ingestion. Treatment is mainly supportive (gastric lavage, activated carbon, fluid resuscitation) but includes various drugs to counter the amatoxins, including intravenous penicillin and cephalosporin derivatives, and, in cases of greater ingestion, can extend to an orthotopic liver transplant. The most reliable method to treat amanitin poisoning is through having the stomach pumped immediately after ingestion; however, the onset of symptoms is generally too late for this to be an option.

Mode of Inhibitory Action
From the crystal structure solved by Dr. Bushnell et. al., α-amanitin interacts with the bridge helix in the RNA polymerase II (pol II). This interaction interferes with the translocation of RNA and DNA needed to empty the site for the next round of RNA synthesis. The addition of α-amanitin can reduce the rate of pol II translocating on DNA from several thousand to a few nucleotides per minute , but has little effect on the affinity of pol II for nucleoside triphosphate, and a phosphodiester bond can still be formed . Bridge helix is designed to be flexibile for its movement is required for translocation. Binding of α-amanitin puts a constraint on its mobility, hence slowing down the synthesis of RNA. People are encouraged to read the original paper published in PNAS (2002, vol. 99, pp1218-1222) for more details.