Disulfide-rich peptide cyclization using sortase A

CI Schroeder, S Kwon and DJ Craik

Institute for Molecular Bioscience, The University of Queensland, Brisbane, 4072, Qld, Australia

Peptides have many advantages over small molecules or antibodies as drug leads in terms of size, high affinity and potency for their physiological targets. Especially disulfide-rich peptides, isolated from plants, scorpions and cone snails, have attracted much interest because of their high specificity for targets including human voltage-gated ion channels, receptors and enzymes as well as their antimicrobial activity. However, linear peptides low stability against proteases in vivo has been a bottleneck for drug development. Therefore, disulfide-rich peptides have been engineered to incorporate a cyclic backbone to improve stability. An attractive alternative method of chemical cyclisation of peptides is enzyme-mediated ligation. Here we describe the use of the bacterial transpeptidase sortase A (SrtA) which catalyses a new peptide bond by recognising a LPXTG sorting motif. We have successfully used SrtA to generate cyclic disulfide-rich peptides of interest for development as cancer therapeutics. Peptides containing one, two, three and four disulfide bonds have rapidly been cyclised in high yield highlighting the versatility of the methodology. Method development, activity, structure and future directions of SrtA-mediated cyclisation will be discussed.