Table 2.

Examples of cyclotides produced using ribosomal expression methods.

Cyclotide	Method	In vitro/in vivo	Pros and Cons	Application	Reference
Kalata B1	Thiol- induced N->S acyl shift	In vitro	Requires GC at ligation site Harsh cyclization conditions	Natural product	(Cowper et al., 2013)
Kalata B1	EPL	Both	Requires N-terminal Cys residues only Yields depend on intein used	Natural product	(R. H. Kimura et al., 2006)
MCoTI-I	EPL	Both	Requires N-terminal Cys residues only Yields depend on intein used	Library scaffold Non-natural amino acid	(Austin et al., 2009; Jagadish et al., 2013)
MCoTI-II	EPL	Both	Requires N-terminal Cys residues only Yields depend on intein used	Library Scaffold	(J. A. Camarero et al., 2007)
MCo-PMI	EPL	in vitro	Requires N-terminal Cys residues only Yields depend on intein used	Anti-cancer	(Ji et al., 2013)
MCoTI-I	PTS	in vivo	Highly efficient (10 times better than EPL) Cyclization can be carried out in Ser/Cys residues Spontaneous self- processing cyclization, no need for enzymes	Library Scaffold Non-natural amino acid	(Jagadish et al., 2013)
rMCoTI-II	SrtA	In vitro	Requires multiple proteolytic steps SrtA recognition sequence of 7 residues remains in the cyclotide after cyclization	Large scale production	(Stanger et al., 2014)