Box 1. Cyclotide synthesis.

Although synthetic macrocycle chemistry has been inspired by the study of natural products such as cyclosporine, the biosynthetic pathways for many compounds remain a mystery. Now, a team led by Professor James Tam at Nanyang Technological University has identified a plant-produced peptide ligase that could be developed as a tool to synthesize cyclic peptides.3

Tam has long studied plants used in traditional Chinese medicine to identify bioactive compounds, and he has previously characterized members from a large and diverse family of plant-produced cyclic peptides known as cyclotides. His team sought to identify the enzyme responsible for cyclotide production in a medicinal plant, Clitoria ternatea.

To identify the peptide ligase that could be responsible for producing the cyclotides in C. ternatea, the team used liquid chromatography and fluorescent reporter constructs to isolate the activity and identify and sequence the protein responsible, which they named butelase 1.

The enzyme was purified and shown to efficiently ligate or cyclize almost any N-terminal residue to a C-terminal asparagine as long as the asparagine is encoded in an asparagine-histidine-valine sequence. The ligase also worked on non-native substrates with the tag appended.

Boston University associate professor Adrian Whitty told SciBX, "The impressive thing about this work is that they were able to find an enzyme that was so broad in its reactivity it requires an asparagine in one of the litigation positions but accepts almost any amino acid in the other, which allows for great freedom of design."

PeptiDream Inc. CSO Patrick Reid said that adapting this technology for peptide-library generation is not trivial, so it is not yet clear whether the ligase will have utility for large-scale compound synthesis.

A first step will be demonstrating that a recombinant form of the enzyme can be expressed and purified.

Patent and licensing information was not available.      -CC