Seven Novel Macrocyclic Polypeptides fromViolaarvensis

Göransson, Ulf; Luijendijk, Teus; Johansson, Senia; Bohlin, Lars; Claeson, Per

doi:10.1021/np9803878

Cited by 165 publications

(125 citation statements)

References 13 publications

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“…Both were susceptible strains to all commercial anthelmintics and had no history of exposure to anthelmintics. Sheep faeces containing eggs of H. contortus or T. colubriformis were sent by overnight courier to Brisbane, the eggs were recovered by passing the faeces through fine sieves (250 m and 75 m), and the sieved material was then centrifuged in a stepwise sucrose gradient (10,25, and 40% sucrose). The eggs were collected from the interface between the 10 and 25% sucrose layers, and washed with water over a 25-m sieve to remove residual sucrose.…”

Section: Methodsmentioning

confidence: 99%

Lysine-scanning Mutagenesis Reveals an Amendable Face of the Cyclotide Kalata B1 for the Optimization of Nematocidal Activity

Huang

Colgrave

Clark

et al. 2010

Journal of Biological Chemistry

108

155

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Cyclotides are a family of macrocyclic peptides that combine the unique features of a head-to-tail cyclic backbone and a cystine knot motif, the combination of which imparts them with extraordinary stability. The prototypic cyclotide kalata B1 is toxic against two economically important gastrointestinal nematode parasites of sheep, Haemonchus contortus and Trichostrongylus colubriformis. A lysine scan was conducted to examine the effect of the incorporation of positive charges into the kalata B1 cyclotide framework. Each of the non-cysteine residues in this 29-amino acid peptide was successively substituted with lysine, and the nematocidal and hemolytic activities of the suite of mutants were determined. Substitution of 11 residues within kalata B1 decreased the nematocidal activity dramatically. On the other hand, six other residues that are clustered on the surface of kalata B1 were tolerant to Lys substitution, and indeed the introduction of positively charged residues into this region increased nematocidal activity. This activity was increased further in double and triple lysine mutants, with a maximal increase (relative to the native kalata B1) of 13-fold obtained with a triple lysine mutant (mutated at positions Thr-20, Asn-29, and Gly-1). Hemolytic activity correlated with the nematocidal activity of all lysine mutants. Our data clearly highlight the residues crucial for nematocidal and hemolytic activity in cyclotides, and demonstrate that the nematocidal activity of cyclotides can be increased by incorporation of basic amino acids.Gastrointestinal nematodes cause major losses to livestock industries worldwide. The control of these pests, until now, has been via synthetic anthelmintics, including benzimidazoles (e.g. thiabendazole), nicotinic acetylcholine receptor agonists (e.g. levamisole), and macrocyclic lactones (e.g. ivermectin and moxidectin). However, resistance of sheep and cattle nematodes to these broad-spectrum anthelmintics is a serious issue for livestock production globally (1-3). Thus, the development of novel agents with potent anthelmintic activity is of great economic and agricultural importance.Cyclotides are circular miniproteins discovered originally in plants of the Rubiaceae, Violaceae, and Cucurbitaceae families (4, 5). Kalata B1, the first cyclotide to be structurally characterized, is abundant in the tropical African plant Oldenlandia affinis (6, 7) but has also been reported in the European Sweet Violet (Viola odorata) (8) and several other Viola species (9 -13). Members of the cyclotide family possess a cyclic peptide backbone and a cystine knot motif, which is formed by three disulfide bonds at the core of their three-dimensional structure. The sequence and structure of kalata B1 is illustrated in Fig. 1, which highlights the six backbone loops between the six conserved cysteine residues that make up the cystine knot (14). The cyclic cystine knot motif is thought to be responsible for the extraordinary enzymatic and chemical stability (15) In a recent study (30), the anthel...

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Section: Methodsmentioning

confidence: 99%

Lysine-scanning Mutagenesis Reveals an Amendable Face of the Cyclotide Kalata B1 for the Optimization of Nematocidal Activity

Huang

Colgrave

Clark

et al. 2010

Journal of Biological Chemistry

108

155

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“…For example, in 1970, kalata B1was reported as the active ingredient in a tea used by women in the Congo region of Africa to accelerate childbirth (16,17), although it was some 25 years later before the sequence and cyclic nature of the peptide were determined (2). The circulins were discovered in screens focusing on anti-HIV activity (3), cyclopsychotride for inhibition of neurotensin binding (4), and various Viola peptides for hemolytic activity (7). Thus, all of the known macrocyclic peptides have diverse biological activities, but their function in plants was not known.…”

mentioning

confidence: 99%

Biosynthesis and insecticidal properties of plant cyclotides: The cyclic knotted proteins from Oldenlandia affinis

Jennings

West

Waine

et al. 2001

Proc. Natl. Acad. Sci. U.S.A.

489

679

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Several members of the Rubiaceae and Violaceae families produce a series of cyclotides or macrocyclic peptides of 29 -31 amino acids with an embedded cystine knot. We aim to understand the mechanism of synthesis of cyclic peptides in plants and have isolated a cDNA clone that encodes the cyclotide kalata B1 as well as three other clones for related cyclotides from the African plant Oldenlandia affinis. The cDNA clones encode prepropeptides with a 20-aa signal sequence, an N-terminal prosequence of 46 -68 amino acids and one, two, or three cyclotide domains separated by regions of about 25 aa. The corresponding cyclotides have been isolated from plant material, indicating that the cyclotide domains are excised and cyclized from all four predicted precursor proteins. The exact processing site is likely to lie on the N-terminal side of the strongly conserved GlyLeuPro or SerLeuPro sequence that flanks both sides of the cyclotide domain. Cyclotides have previously been assigned an antimicrobial function; here we describe a potent inhibitory effect on the growth and development of larvae from the Lepidopteran species Helicoverpa punctigera.

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“…Soon after, a series of peptides from viola plants was identified (1,8), and the various peptides discovered to that point were recognized as part of a protein family that was named the cyclotides (1). Additional members have been discovered in the last few years (9 -14).…”

mentioning

confidence: 99%

Twists, Knots, and Rings in Proteins

Rosengren

Daly

Plan

et al. 2003

Journal of Biological Chemistry

299

285

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In recent years an increasing number of miniproteins containing an amide-cyclized backbone have been discovered. The cyclotide family is the largest group of such proteins and is characterized by a circular protein backbone and six conserved cysteine residues linked by disulfide bonds in a tight core of the molecule. These form a cystine knot in which an embedded ring formed by two of the disulfide bonds and the connecting backbone segment is threaded by a third disulfide bond. In the current study we have undertaken high resolution structural analysis of two prototypic cyclotides, kalata B1 and cycloviolacin O1, to define the role of the conserved residues in the sequence. We provide the first comprehensive analysis of the topological features in this unique family of proteins, namely rings (a circular backbone), twists (a cis-peptide bond in the Mö bius cyclotides) and knots (a knotted arrangement of the disulfide bonds).

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Seven Novel Macrocyclic Polypeptides fromViolaarvensis

Cited by 165 publications

References 13 publications

Lysine-scanning Mutagenesis Reveals an Amendable Face of the Cyclotide Kalata B1 for the Optimization of Nematocidal Activity

Lysine-scanning Mutagenesis Reveals an Amendable Face of the Cyclotide Kalata B1 for the Optimization of Nematocidal Activity

Biosynthesis and insecticidal properties of plant cyclotides: The cyclic knotted proteins from Oldenlandia affinis

Twists, Knots, and Rings in Proteins

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