Peptidomics of Circular Cysteine-Rich Plant Peptides: Analysis of the Diversity of Cyclotides from Viola tricolor by Transcriptome and Proteome Mining

J Proteome Res. 2015 Nov 6;14(11):4851-62. doi: 10.1021/acs.jproteome.5b00681. Epub 2015 Oct 8.


Cyclotides are plant-derived mini proteins. They are genetically encoded as precursor proteins that become post-translationally modified to yield circular cystine-knotted molecules. Because of this structural topology cyclotides resist enzymatic degradation in biological fluids, and hence they are considered as promising lead molecules for pharmaceutical applications. Despite ongoing efforts to discover novel cyclotides and analyze their biodiversity, it is not clear how many individual peptides a single plant specimen can express. Therefore, we investigated the transcriptome and cyclotide peptidome of Viola tricolor. Transcriptome mining enabled the characterization of cyclotide precursor architecture and processing sites important for biosynthesis of mature peptides. The cyclotide peptidome was explored by mass spectrometry and bottom-up proteomics using the extracted peptide sequences as queries for database searching. In total 164 cyclotides were discovered by nucleic acid and peptide analysis in V. tricolor. Therefore, violaceous plants at a global scale may be the source to as many as 150 000 individual cyclotides. Encompassing the diversity of V. tricolor as a combinatorial library of bioactive peptides, this commercially available medicinal herb may be a suitable starting point for future bioactivity-guided screening studies.

Keywords: 1kp; Violaceae; circular peptides; cystine-knot; mass spectrometry; natural products; peptidomics; ribosomally synthesized and post-translationally modified peptides; transcriptome.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Chromatography, High Pressure Liquid
  • Cyclotides / chemistry*
  • Cyclotides / genetics
  • Cyclotides / isolation & purification
  • Cyclotides / metabolism
  • Cystine Knot Motifs / genetics
  • Data Mining
  • Gene Expression Regulation, Plant*
  • Gene Library
  • Liquid-Liquid Extraction
  • Models, Molecular
  • Molecular Sequence Data
  • Plant Components, Aerial / chemistry
  • Plant Extracts / chemistry
  • Plant Proteins / chemistry
  • Plant Proteins / genetics*
  • Plant Proteins / isolation & purification
  • Plant Proteins / metabolism
  • Protein Processing, Post-Translational*
  • Proteome / genetics
  • Proteome / metabolism
  • Proteomics / methods
  • Sequence Alignment
  • Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
  • Transcriptome*
  • Violaceae / genetics*
  • Violaceae / metabolism


  • Cyclotides
  • Plant Extracts
  • Plant Proteins
  • Proteome