Cyclotides from an Extreme Habitat: Characterization of Cyclic Peptides from Viola abyssinica of the Ethiopian Highlands

Abstract

As part of ongoing explorations of the structural diversity of cyclotides, the cyclotide content of a native violet of the East African highlands, Viola abyssinica (which grows at altitudes up to 3400 m), was studied. Six new cyclotides, vaby A-E (1-5) and varv E (6), were isolated and characterized by employing HPLC and MS techniques and quantitative amino acid analysis. Cyclotides 1-5 were found to have new sequences, and 1-3 have a further novel feature in their sequences, an alanine moiety in loop 2. Two of the cyclotides (1 and 4) also exhibited cytotoxic properties in a flourometric microculture cytotoxicity assay. The findings corroborate the hypothesis that investigating the cyclotide contents of violets growing in diverse environments is a promising approach for extending our knowledge of both the structural and biological diversity of cyclotides.

Cyclotide Structure–Activity Relationships: Qualitative and Quantitative Approaches Linking Cytotoxic and Anthelmintic Activity to the Clustering of Physicochemical Forces

“…We show that these descriptors are useful for characterizing and classifying cyclotides, and for understanding their structure-activity relationships. To this end, we analyze the physicochemical properties of the 68 known cyclotides with cytotoxic activity [13], [18], [22], [47], [48], [49], [50] against the lymphoma cell line U937-GTB or anthelmintic activity [20], [51] against the caterpillar larvae of H. contortus .…”

“…Cytotoxic [13], [18], [22], [47], [48], [49], [50] and anthelmintic [20], [51] activity data were obtained from the literature. Table 2 summarizes the potency of cyclotides and shows their IC 50 values.…”

Anionic Host Defence Peptides from the Plant Kingdom: Their Anticancer Activity and Mechanisms of Action

“…In contrast, vaby D (net charge -1) was isolated from Viola abyssinica which grows in extreme habitats and altitudes in the Ethiopian Highlands. The peptide was found to have a broad spectrum of anticancer activity with IC 50 values ranging between 3 μM to 47 μM when directed against a variety of cancers, including hepatocellular carcinoma cells (BEL-7402), and breast carcinoma (MDA-MB-231) [42,73]. More recently, the mechanisms underpinning the activity of the peptide to U-937 GTB lymphoma cells was investigated, which suggested that, in addition to targeting the membranes of these cells, the peptide may be taken up to inflict damage on DNA and other internal targets [74].…”

The bountiful biological activities of cyclotides

“…[44] The use of cyclotides in human health applications was first explored during the discovery of kalata B1, and although its uterotonic activity was established in rat, rabbit, and human uteri, it is not recommended as an oxytocic agent because of the severe side effects. [4] During the past decade, a profusion of bioactivity-directed research demonstrates that cyclotides display an assortment of activities, including antineurotensin, [10] trypsin inhibiting, [22,23] hemolytic, [30,31] cytotoxic/antitumor, [45][46][47][48][49][50][51][52][53][54][55][56] and anti-HIV activities; [9,29] several of these properties have prospective therapeutic relevance.…”

“…Psyle A -F U-937GTB, MCF-7, MCF-7/ADR 0.72 to >10 [51,52] Viola odorata CyO2 A549, ACHN, BEL-7402, BGC-823, BEL-7402, DU145, CCRF-CEM, CCRF-CEM/VM-1, CLL,NCI-H66, NCI-H69AR, MDA-MB-231, OVCA, PBMC, RPMI-8226/s, RPMI-8226/ Dox40, RPMI-8226/LR-5, U251, U-937GTB, U-937VcR 0.1 to 17.05 [45,49,52,55] V. abyssinica Vaby A U-937 GTB 7.6 [56] Vaby D A549, BEL-7402, BGC-823, DU145, MDA-MB-231, U251, U-937 GTB 2.8 to 46.62 [53,56] V. arvensis Varv A ACHN, CCRF-CEM, CCRF-CEM/VM-1, CLL,NCI-H66, NCI-H69AR, OVCA, PBMC, RPMI-8226/s, RPMI-8226/ Dox40, RPMI-8226/LR-5, U-937GTB, U-937VcR 2.7 to 12.1 [45] Varv F ACHN, CCRF-CEM, CCRF-CEM/VM-1, CLL,NCI-H66, NCI-H69AR, OVCA, PBMC, RPMI-8226/s, RPMI-8226/ Dox40, RPMI-8226/LR-5, U-937GTB, U-937VcR 2.6 to 7.4 [45] V. labridorica Vila A A549, BEL-7402, BGC-823, DU145, MDA-MS-123, U251 5.08 to >10 [49] Vila B A549, BEL-7402, BGC-823, DU145, MDA-MS-123, U251 6.25 to 34.65 [49] Vila D A549, BEL-7402, BGC-823, DU145, MDA-MS-123, U251 >10 to 49.59 [49] V. philippica Viphi A-G BGC-823, HeLa, HFF-1, MM96L 1.03 to 6.35 [55] Viba 15, 17, Varv A, Kalata B1 BGC-823, HeLa, HFF-1, MM96L 1.32 to 10.21 [55] V. tricolor Varv A,D, E, F, H, He, Hm RPMI-8226/s, U-937GTB 4 to 74.39 [46,54] Vitri A-F A549, BEL-7402, DU145, MDA-MB-231, RPMI-8226/s, U251, U-937GTB 0.6 to 54.39 [46,54] V. biflora Vibi D U-937GTB >30 [49] Vibi E U-937GTB 3.2 [49] Vibi G U-937GTB 1 [49] Vibi H U-937GTB 1.6 [49] A549 Two amino acid residues that can impact potency of several bioactivities are the conserved Glu in loop 1 and tryptophan (Trp, W). For instance, the esterification of Glu in the bracelet cyclotide, CyO 2 , results in a 48-fold decrease in cytotoxic potency [48] and a near loss of antibacterial activity against Salmonella sp.…”