More details about the structure of HC-toxin

Cyclic peptides (more generally and correctly known as non-ribosomally synthesized peptides or NRPs) are an important and widespread class of secondary metabolites. Many of them are biologically active and well-known for their medical and pharmaceutical properties. For example, penicillin, and the immunosuppressant cyclosporin are all non-ribosomally synthesized peptides (Fig. 1). 

Examples of nonribosomal peptides (NRPs) from fungi.

Fig. 1 Examples of nonribosomal peptides (NRPs) from fungi. From Walton et al., (2004) Peptide synthesis without ribosomes. In: Advances in Fungal Biotechnology for Industry, Agriculture, and Medicine. J. Tkacz and L. Lange, eds., Kluwer Academic, New York, pp. 127-162. (NB: Originally we thought that amatoxins from poisonous Amanita mushrooms are also NRPs, but we were wrong. They are ribosomally encoded; see Hallen et al., 2007, PNAS 104:19097). A: destruxin. B: amanitin. C: cyclosporin. D: ergotamine. E: HC-toxin. F: AM-toxin. G: ferrichrome. H: enniatin. I: victorin. J: ACV. K: peptaibol.

NRPs are biosynthesized by a family of functionally and structurally related enzymes known as non-ribosomal peptide synthetases (NRPSs). (Peptides such as glutathione are made by an unrelated biochemical pathway involving sequential independent enzymatic steps).

For example, the first committed step in penicillin biosynthesis is catalyzed by "ACV synthetase". This enzyme uses L-amino acids to make the linear tripeptide "L-aminoadipyl-L-cysteinyl-D-valine". This compound is then rearranged by another enzyme to make the beta-lactam ring, and then typically further modified to make the family of penicillins and related cephalosporins.

Cyclic peptides often contain D-amino acids. In some cases the L amino acid is converted to the D form by the NRPS itself. For example, ACV synthetase has an 'epimerase' domain in its third domain that converts L-valine to D-valine. HC-toxin contains two D amino acids. The first one, D-proline, is made from L-proline by an epimerase domain within module1 of HC-toxin synthetase. However, the second D amino acid, D-alanine, is made by a separate enzyme (alanine racemase, the product of TOXG; Cheng and Walton, 2000). The D-alanine is then activated by the third module of HC-toxin synthetase for incorporation into HC-toxin.

Cyclic peptides are cunning molecules. For example, they can adopt different conformations, i.e., different stereochemistries of their peptide bonds, depending on their solvent environment. Some conformers of some cyclic peptides interconvert so slowly that they can be resolved by chromatography. In chloroform, HC-toxin has one stable conformer whereas in water it is a mixture of two conformers. Its solubility in both hydrophilic and hydrophoblic solvents is probably important for the biological activity of HC-toxin, allowing it to readily penetrate plant tissues and cells.

The side chain of Aeo is like a scorpion's tail. Both the terminal epoxide and the vicinal carbonyl group are necessary for activity. (However, apicidin does not have the epoxide but is still an HDAC inhibitor; Jin et al., Mol. Microbiol. 76:456, 2010). The proline helps the peptide cyclize against the strain of the four peptide bonds in a tight ring. The alanine residues are probably not critical since other cyclic peptides with substitutions at these positions still have the same biological activity.

Cyclic tetrapeptides containing Aeo (or the related 2-amino decanoic acid) are made by at least five filamentous fungi. An intriguing question is whether this is situation is due to convergent evolution, to common descent from an ancestor that makes Aeo-containing tetrapeptides, or to horizontal gene transfer.

For the structures of the other Aeo-containing peptides, go to this page.