Our technology will bring new medicines for diseases of unmet clinical need
We have created a chemoenzymatic technology for macrocycle generation. The technology consists of a set of flexible engineered enzymes which can carry out modification steps on a synthetic linear substrate to generate modified macrocycles.
The biosynthetic enzymes originate from a number of different freshwater and marine symbiotic or free living cyanobacteria (blue-green algae) and are responsible for the production of a family of ribosomally produced and post-translationally modified peptides (RiPPs) called the cyanobactins. Cyanobactins are a large group of macrocycles which show diversity in both ring size and chemistry. The extensive decoration of the ring has many attractive features for drug design (Fig. 1).
These macrocyclic peptides are defined by a C to N terminal peptide bond, the incorporation of heterocycles at different oxidation states (Thiazoline ThH, Oxazoline OxH, Thiazole Thz, Oxazole Oxz) and possible decoration with prenyl groups. Some cyanobactins incorporate inverted stereocentres at the Cα adjacent to ThH/Thz.
We have used our knowledge of the structures and mechanisms of the individual enzymes of a number of cyanobactin biosynthesis pathways to engineer them so that they can be used for fast, cost-effective synthesis of a range of modified macrocycles from exogenously-supplied linear peptide substrates. Our modified heterocyclase (the PatE gene product), has been engineered to be able to process a range of substrates without the need for a long leader peptide (Fig. 2). This provides for increased reaction mass efficiency. The modified enzyme is able to create a number of different rings containing hetero-atoms (Fig. 3).
We have broadened the enzymes' substrate specificity, as well as removing the need for a large N-terminal recognition sequence (leader) in the substrate peptide. The substrate can be 6-12 residues in length and may contain L- and/or D-amino acids and residues with non-natural side groups. The engineered enzymes can already be produced at gram scale and work at appreciable rates. The product can be produced at multi-milligram scale with larger scales possible.
Novel and diverse GyrocycleTM macrocyclic peptides can be created by a combination of biotransformation and chemical steps (see Fig. 4).