Cyclotides are naturally occurring plant-based proteins of approximately 30 amino acids in size that contain a head-to-tail cyclized backbone and a cystine knot motif formed by their three conserved disulfide bonds. Their exceptional stability and unique topology make them valuable frameworks in drug design or protein engineering applications. To facilitate such applications and to explore structure–activity relationships of cyclotides it is useful to be able to chemically synthesize them, a process that is readily achieved via solid phase peptide synthesis followed by oxidative folding. This chapter describes what is known about the oxidative folding of cyclotides, both in chemical folding buffers and assisted by a protein disulfide isomerase enzyme isolated from a cyclotide-producing plant. Formation of the cystine knot motif is readily achieved, despite its apparent topological complexity.