Isolation, Characterization, and Bioactivity of Cyclotides from the Micronesian Plant Psychotria leptothyrsa

Abstract

Cyclotides, the largest known family of head-to-tail cyclic peptides, have approximately 30 amino acid residues with a complex structure containing a circular peptide backbone and a cystine knot. They are found in plants from the Violaceae and Rubiaceae families and are speculated to function in plant protection. In addition to their insecticidal properties, cyclotides display cytotoxic, anti-HIV, antimicrobial, and inhibition of neurotensin binding activities. Although cyclotides are present in all violaceous species hitherto screened, their distribution and expression in Rubiaceae are not fully understood. In this study, we show that Psychotria leptothyrsa var. longicarpa (Rubiaceae) contains a suite of different cyclotides. The cyclotide fractions were isolated by RP-HPLC, and sequences of six new peptides, named psyles A-F, were determined by MS/MS sequencing. One of these, psyle C, is the first rubiaceous linear variant known. Psyles A, C, and E were analyzed in a fluorometric microculture assay to determine cytotoxicity toward the human lymphoma cell line U937-GTB. The IC(50) values of psyles A, C, and E were 26, 3.50, and 0.76 muM, respectively. This study expands the number of known rubiaceous cyclotides and shows that the linear cyclotide maintains cytotoxicity.

Discovery of a Linear Cyclotide from the Bracelet Subfamily and Its Disulfide Mapping by Top-down Mass Spectrometry

“…Hedyotide B2, surprisingly, has a linear structure because it lacks the essential Asn/Asp residue in the putative loop 6 of the cyclized form. Although few examples of linear cyclotides have been identified, they all belong to the Möbius subfamily (21)(22)(23). This makes hedyotide B2 the first linear variant or uncyclotide of the bracelet subfamily to be discovered.…”

“…Recently, a few linear variants of cyclotides have been discovered (21,22). Their occurrences appear to be rare, with only two naturally occurring linear variants having been reported: violacin A from Viola odorata (21) and psyle C from Psychotria leptothyrsa (22).…”

“…Their occurrences appear to be rare, with only two naturally occurring linear variants having been reported: violacin A from Viola odorata (21) and psyle C from Psychotria leptothyrsa (22). Two other linear cyclotides have been described from O. affinis but could have been linearized from their cyclic forms by the chemical degradation of the Asn-Gly or Asp-Gly bond of loop 6 through the aspartimide formation and subsequent ring opening during the isolation procedure or the decomposition of dried plant samples (23).…”

Discovery of Linear Cyclotides in Monocot Plant Panicum laxum of Poaceae Family Provides New Insights into Evolution and Distribution of Cyclotides in Plants

“…Both mutations can account for the failure in the biosynthesis of cyclotide-containing precursors to produce the characteristic cyclic peptide backbone of cyclotides. Although the functional correlation between the circular structure and bioactivity remains controversial, several linear cyclotides have been shown to retain the bioactivities comparable with their cyclic counter parts (18,23).…”

Novel Cyclotides and Uncyclotides with Highly Shortened Precursors from Chassalia chartacea and Effects of Methionine Oxidation on Bioactivities

“…Recently, a few linear variants of cyclotides (also known as uncyclotides) have been identified, including violacin A from Viola odorata (23), psyle C from Psychotria leptothyrsa (24), and hedyotide B2 from Hedyotis biflora (22). Genetic characterization showed that both violacin A and hedyotide B2 have similar biosynthetic processing (22,23).…”