Cyclotides are a unique class of cyclic peptides exclusive to the plant kingdom, rich in cysteine, consisting of 28 to 37 amino acids. Their special head-to-tail cyclic cystine knot (CCK) structural motif imbues cyclotides with remarkable stability, a quality that makes them promising molecules for diverse applications in agriculture and pharmaceuticals.

Cyclotides have been discovered in five major plant families: Violaceae, Rubiaceae, Cucurbitaceae, Solanaceae, and Fabaceae. Among these, the Violaceae family is particularly abundant in cyclotides; indeed, every examined plant within this family has been found to contain these cyclic peptides.

In the current study, the occurrence of cyclotides in Viola communis was investigated, and 20 cyclotides were isolated at the protein level including two new cyclotides (Vcom1 and Vcom2). The structural analysis for the newly isolated peptides demonstrated a similar structural fold to the previously characterized cyclotides. Additionally, the insecticidal activity of the newly isolated cyclotides was tested to evaluate their efficacy against the target insects.

Transcriptome analyses of Viola odorata and V. communis were performed for the first time. Seven new cyclotide-like sequences were observed in the transcriptome analysis of V. communis. Here, we attempt to investigate the multimeric sequences and precursor’s structure in these plants and by examining the transcriptomes of these two species side by side, we endeavor to understand the potential variations in key biological processes including the cyclization process and cyclotide production patterns.