Backbone Dynamics of Cyclotide MCoTI‐I Free and Complexed with Trypsin

Puttamadappa, Shadakshara S.; Jagadish, Krishnappa; Shekhtman, Alexander; Camarero, Julio A.

doi:10.1002/anie.201002906

Cited by 56 publications

(67 citation statements)

References 36 publications

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“…The interlocking nature of cyclic cystine knot (CCK) motif makes the cyclotide backbone very compact providing a highly rigid structure. [33] As a result, cyclotides are extremely stable compounds, that have been shown to be resistant to thermal and chemical denaturation and to enzymatic degradation. [17a, 34] Proof of this extraordinary stability is evident from the discovery of the first cyclotide, kalata B1, which was able to remain structurally intact and biologically active after being extracted by boiling water to make a medicinal herbal tea.…”

Section: Structurementioning

confidence: 99%

Cyclotides: Overview and Biotechnological Applications

Gould

Camarero

2017

ChemBioChem

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Cyclotides are globular microproteins with a unique head-to-tail cyclized backbone, stabilized by three disulfide bonds forming a cystine knot. This unique circular backbone topology and knotted arrangement of three disulfide bonds makes them exceptionally stable to chemical, thermal and biological degradation compared to other peptides of similar size. In addition, cyclotides have been shown to be highly tolerant to sequence variability aside from the conserved residues forming the cystine knot. Cyclotides can also cross cellular membranes and are able to modulate intracellular protein-protein interactions both in vitro and in vivo. All these features make cyclotides appear as highly promising leads or frameworks for the design of peptide-based diagnostic and therapeutic tools. This article provides an overview on cyclotides and their applications as molecular imaging agents and peptide-based therapeutics.

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Section: Structurementioning

confidence: 99%

Cyclotides: Overview and Biotechnological Applications

Gould

Camarero

2017

ChemBioChem

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“…Overall, the order parameters of HTD-2 are in agreement with those reported for two other cyclic disulfide-rich peptides: MCoTI-I and MCoTI-II. Specifically, HTD-2 has S 2 values of 0.67–0.96, similar to ranges of 0.76–0.94 and 0.72–0.91 for MCoTI-I(39) and MCoTI-II,(10) respectively. The proposed “butterfly”-type motion of the θ-defensin structure cannot be verified by the differences in the S 2 parameter between the β-strand and turn regions because flexibility described by the S 2 parameter is on the picosecond–nanosecond time scale, whereas a “butterfly”-type bending would occur on the millisecond time scale.…”

Section: Resultsmentioning

confidence: 69%

“…In the case of the cyclic cystine knot proteins, for example, MCoTI-I and MCoTI-II, variations in the flexibility of the binding site have been reported between bound and free forms. (10, 39, 41)…”

Section: Resultsmentioning

confidence: 99%

Insights into the Molecular Flexibility of θ-Defensins by NMR Relaxation Analysis

Conibear¹,

Wang²,

et al. 2014

J. Phys. Chem. B

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θ-Defensins are mammalian cyclic peptides that have antimicrobial activity and show potential as stable scaffolds for peptide-based drug design. The cyclic cystine ladder structural motif of θ-defensins has been characterized using NMR spectroscopy and is important for their structure and stability. However, the effect of the pronounced elongated topology of θ-defensins on their molecular motion is not yet understood. Studies of molecular motion by NMR relaxation measurements have been facilitated by the recent development of a semirecombinant method for producing cyclic peptides that allows for isotopic labeling. Here we have undertaken a multifield 15N NMR relaxation analysis of the anti-HIV θ-defensin, HTD-2, and interpreted the experimental data using various models of overall and internal molecular motion. We found that it was necessary to apply a model that includes internal motion to account for the variations in the experimental T1 and NOE data at different backbone amide sites in the peptide. Although an isotropic model with internal motion was the simplest model that provided a satisfactory fit with the experimental data, we cannot exclude the possibility that overall motion is anisotropic, especially considering the strikingly elongated topology of θ-defensins. The presence of flexible side chains, self-association, interactions with solvent, and internal motions are all potential contributors to the observed relaxation data. Internal motion consistent with the constraints imposed by the cyclic cystine ladder was observed in that the order parameters, S2, show that residues in the turns are more flexible than those in the β-sheet. This study provides insights into the dynamics of θ-defensins and information that might be useful in their application as scaffolds in drug design.

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“…[24] AC P4-derived linear peptide,inwhich the Cys residue was replaced by Ser,was grafted onto the cyclotide scaffold using loop 6t op rovide cyclotide MCoCP4 (Figure 1). We have shown that loop 6h as the higher backbone mobility of the MCoTI-I cyclotide [3] and accordingly this loop has been shown to tolerate well the grafting of bioactive peptides. [6a, 7] Replacement of the Cys residue was done to facilitate the folding of MCoCP4.…”

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confidence: 99%

“…[2] They share au nique head-to-tail circular knotted topology of three disulfide bridges,with one disulfide penetrating through amacrocycle formed by the two other disulfides and interconnecting peptide backbones, forming what is called ac ystine knot topology ( Figure 1A). This cyclic cystine knot (CCK) framework gives the cyclotides exceptional rigidity, [3] resistance to thermal and chemical denaturation, and enzymatic stability against degradation. [2] Interestingly,s ome cyclotides have been shown to be orally bioavailable, [4] and other cyclotides have been shown to cross the cell membrane through macropinocytosis.…”

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confidence: 99%

Recombinant Expression and Phenotypic Screening of a Bioactive Cyclotide Against α‐Synuclein‐Induced Cytotoxicity in Baker′s Yeast

et al. 2015

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We report for the first time the recombinant expression of fully folded bioactive cyclotides inside live yeast cells by using intracellular protein trans-splicing in combination with ahighly efficient split-intein. This approach was successfully used to produce the naturally occurring cyclotide MCoTI-I and the engineered bioactive cyclotide MCoCP4. Cyclotide MCoCP4 was shown to reduce the toxicity of human a-synuclein in live yeast cells.C yclotide MCoCP4 was selected by phenotypic screening from cells transformed with am ixture of plasmids encoding MCoCP4 and inactive cyclotide MCoTI-I in ar atio of 1:510 4 .T his demonstrates the potential for using yeast to perform phenotypic screening of genetically encoded cyclotide-based libraries in eukaryotic cells.

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Backbone Dynamics of Cyclotide MCoTI‐I Free and Complexed with Trypsin

Cited by 56 publications

References 36 publications

Cyclotides: Overview and Biotechnological Applications

Cyclotides: Overview and Biotechnological Applications

Insights into the Molecular Flexibility of θ-Defensins by NMR Relaxation Analysis

Recombinant Expression and Phenotypic Screening of a Bioactive Cyclotide Against α‐Synuclein‐Induced Cytotoxicity in Baker′s Yeast

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