Ribosomal biosynthesis of alpha-amanitin and related cyclic peptides from agarics

The toxins of deadly poisonous Amanita mushrooms, such as α-amanitin and phalloidin, are ribosomally synthesized cyclic peptides encoded by the “MSDIN” gene family. The members of this family encode 33- to 37-amino acid propeptides with conserved upstream and downstream regions flanking a highly variable core region of 6-10 amino acids. The full MSDIN family has been characterized in two amanitin-producing mushrooms, the “death cap” A. phalloides and the “destroying angel” A. bisporigera. Each species has ~30 MSDIN genes. Only three genes (α-amanitin, phallacidin, and an unknown) are common to both species; between them they have the capacity to encode ~56 unique cyclic peptides. One of the A. phalloides genes encodes cycloamanide B, a chemically known immunosuppressive homodetic cyclic heptapeptide, but genes for the other known cycloamanides were not found in the genome. Based on their predicted masses, three new cycloamanides (E-F) were detected in chloroform extracts of A. phalloides. Both A. phalloides and A. bisporigera contain two prolyl oligopeptidase genes, one of which (POPB) is dedicated to cyclic peptide biosynthesis. POPB processes the 35-amino acid propeptide of α-amantin in two steps: proteolysis of the 10-amino acid leader peptide and nonprocessive transpeptidation of the core amino acids to produce the cyclic octapeptide cyclo(IWGIGCNP). POPB is a versatile peptide cyclase that can cyclize a broad range of synthetic peptides. Galerina marginata and Lepiota subincarnata, two other poisonous agarics, also produce α-amanitin ribosomally, but the propeptide sequences are not well-conserved outside their core regions.