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David J. Craik - One of the best experts on this subject based on the ideXlab platform.
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co expression of a cyclizing asparaginyl endopeptidase enables efficient production of Cyclic Peptides in planta
Journal of Experimental Botany, 2018Co-Authors: Simon Poon, David J. Craik, Thomas Durek, Karen S Harris, Edward K Gilding, Nicole L Van Der Weerden, Mark A Jackson, Owen C Mccorkelle, Marilyn A AndersonAbstract:Cyclotides are ultra-stable, backbone-cyclized plant defence Peptides that have attracted considerable interest in the pharmaceutical industry. This is due to their range of native bioactivities as well as their ability to stabilize other bioactive Peptides within their framework. However, a hindrance to their widespread application is the lack of scalable, cost-effective production strategies. Plant-based production is an attractive, benign option since all biosynthetic steps are performed in planta. Nonetheless, cyclization in non-cyclotide-producing plants is poor. Here, we show that Cyclic Peptides can be produced efficiently in Nicotiana benthamiana, one of the leading plant-based protein production platforms, by co-expressing cyclotide precursors with asparaginyl endopeptidases that catalyse peptide backbone cyclization. This approach was successful in a range of other plants (tobacco, bush bean, lettuce, and canola), either transiently or stably expressed, and was applicable to both native and engineered Cyclic Peptides. We also describe the use of the transgenic system to rapidly identify new asparaginyl endopeptidase cyclases and interrogate their substrate sequence requirements. Our results pave the way for exploiting cyclotides for pest protection in transgenic crops as well as large-scale production of Cyclic peptide pharmaceuticals in plants.
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Molecular Grafting onto a Stable Framework Yields Novel Cyclic Peptides for the Treatment of Multiple Sclerosis
2016Co-Authors: Natalie Payne, Guizhi Sun, Shunhe Wang, Claude C. Bernard, David J. CraikAbstract:ABSTRACT: Multiple sclerosis (MS) is an inflammatory disease of the central nervous system (CNS) and is characterized by the destruction of myelin and axons leading to progressive disability. Peptide epitopes from CNS proteins, such as myelin oligodendrocyte glycoprotein (MOG), possess promising immunoregulatory potential for treating MS; however, their instability and poor bioavailability is a major impediment for their use clinically. To overcome this problem, we used molecular grafting to incorporate peptide sequences from the MOG35−55 epitope onto a cyclotide, which is a macroCyclic peptide scaffold that has been shown to be intrinsically stable. Using this approach, we designed novel Cyclic Peptides that retained the structure and stability of the parent scaffold. One of the grafted Peptides, MOG3, displayed potent ability to prevent disease development in a mouse model of MS. These results demonstrate the potential of bioengineered Cyclic Peptides for the treatment of MS. Multiple sclerosis (MS) is an inflammatory disorder of thecentral nervous system (CNS) characterized by focal demyelinating lesions,1 where both the cellular and humora
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Molecular Grafting onto a Stable Framework Yields Novel Cyclic Peptides for the Treatment of Multiple Sclerosis
2015Co-Authors: Conan K. Wang, Christian W. Gruber, Maša Cemazar, Christopher Siatskas, Prascilla Tagore, Natalie Payne, Guizhi Sun, Shunhe Wang, Claude C. Bernard, David J. CraikAbstract:Multiple sclerosis (MS) is an inflammatory disease of the central nervous system (CNS) and is characterized by the destruction of myelin and axons leading to progressive disability. Peptide epitopes from CNS proteins, such as myelin oligodendrocyte glycoprotein (MOG), possess promising immunoregulatory potential for treating MS; however, their instability and poor bioavailability is a major impediment for their use clinically. To overcome this problem, we used molecular grafting to incorporate peptide sequences from the MOG35–55 epitope onto a cyclotide, which is a macroCyclic peptide scaffold that has been shown to be intrinsically stable. Using this approach, we designed novel Cyclic Peptides that retained the structure and stability of the parent scaffold. One of the grafted Peptides, MOG3, displayed potent ability to prevent disease development in a mouse model of MS. These results demonstrate the potential of bioengineered Cyclic Peptides for the treatment of MS
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translational diffusion of Cyclic Peptides measured using pulsed field gradient nmr
Journal of Physical Chemistry B, 2014Co-Authors: Conan K Wang, Susan E Northfield, Joakim E Swedberg, Peta J Harvey, Alan M Mathiowetz, D Price, Spiros Liras, David J. CraikAbstract:Cyclic Peptides are increasingly being recognized as valuable templates for drug discovery or design. To facilitate efforts in the structural characterization of Cyclic Peptides, we explore the use of pulse-field gradient experiments as a convenient and noninvasive approach for characterizing their diffusion properties in solution. We present diffusion coefficient measurements of five Cyclic Peptides, including dichC, SFTI-1, cVc1.1, kB1, and kB2. These Peptides range in size from six to 29 amino acids and have various therapeutically interesting activities. We explore the use of internal standards, such as dioxane and acetonitrile, to evaluate the hydrodynamic radius from the diffusion coefficient, and show that 2,2-dimethyl-2-silapentane-5-sulfonic acid, a commonly used chemical shift reference, can be used as an internal standard to avoid spectral overlap issues and simplify data analysis. The experimentally measured hydrodynamic radii correlate with increasing molecular weight and in silico predictions. We further applied diffusion measurements to characterize the self-association of kB2 and showed that it forms oligomers in a concentration-dependent manner, which may be relevant to its mechanism of action. Diffusion coefficient measurements appear to have broad utility in Cyclic peptide structural biology, allowing for the rapid characterization of their molecular shape in solution.
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Cyclic Peptides arising by evolutionary parallelism via asparaginyl endopeptidase mediated biosynthesis
The Plant Cell, 2012Co-Authors: Joshua S Mylne, Lai Yue Chan, Aurelie H Chanson, Norelle L Daly, Hanno Schaefer, Timothy L Bailey, Philip Nguyencong, Laura Cascales, David J. CraikAbstract:The Cyclic miniprotein Momordica cochinchinensis Trypsin Inhibitor II (MCoTI-II) (34 amino acids) is a potent trypsin inhibitor (TI) and a favored scaffold for drug design. We have cloned the corresponding genes and determined that each precursor protein contains a tandem series of Cyclic TIs terminating with the more commonly known, and potentially ancestral, aCyclic TI. Expression of the precursor protein in Arabidopsis thaliana showed that production of the Cyclic TIs, but not the terminal aCyclic TI, depends on asparaginyl endopeptidase (AEP) for maturation. The nature of their repetitive sequences and the almost identical structures of emerging TIs suggest these Cyclic Peptides evolved by internal gene amplification associated with recruitment of AEP for processing between domain repeats. This is the third example of similar AEP-mediated processing of a class of Cyclic Peptides from unrelated precursor proteins in phylogenetically distant plant families. This suggests that production of Cyclic Peptides in angiosperms has evolved in parallel using AEP as a constraining evolutionary channel. We believe this is evolutionary evidence that, in addition to its known roles in proteolysis, AEP is especially suited to performing protein cyclization.
Peter Proksch - One of the best experts on this subject based on the ideXlab platform.
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callyaerins from the marine sponge callyspongia aerizusa Cyclic Peptides with antitubercular activity
Journal of Natural Products, 2015Co-Authors: Georgios Daletos, Victor Wray, Wenhan Lin, Rainer Kalscheuer, Rudolf Hartmann, Nicole J. De Voogd, Hendrik Koliwerbrandl, Peter ProkschAbstract:Chemical investigation of the Indonesian sponge Callyspongia aerizusa afforded five new Cyclic Peptides, callyaerins I-M (1-5), along with the known callyaerins A-G (6-12). The structures of the new compounds were unambiguously elucidated on the basis of one- and two-dimensional NMR spectroscopy and mass spectrometry. In addition, the structures of callyaerins D (9), F (11), and G (12), previously available in only small amounts, have been reinvestigated and revised. All compounds were tested in vitro against Mycobacterium tuberculosis, as well as against THP-1 (human acute monocytic leukemia) and MRC-5 (human fetal lung fibroblast) cell lines, in order to assess their general cytotoxicity. Callyaerins A (6) and B (7) showed potent anti-TB activity with MIC₉₀ values of 2 and 5 μM, respectively. Callyaerin C (8) was found to be less active, with an MIC₉₀ value of 40 μM. Callyaerin A (6), which showed the strongest anti-TB activity, was not cytotoxic to THP-1 or MRC-5 cells (IC₅₀ > 10 μM), which highlights the potential of these compounds as promising anti-TB agents.
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Callyaerins from the Marine Sponge Callyspongia aerizusa: Cyclic Peptides with Antitubercular Activity
2015Co-Authors: Georgios Daletos, Victor Wray, Wenhan Lin, Rainer Kalscheuer, Hendrik Koliwer-brandl, Rudolf Hartmann, Nicole J. De Voogd, Peter ProkschAbstract:Chemical investigation of the Indonesian sponge Callyspongia aerizusa afforded five new Cyclic Peptides, callyaerins I–M (1–5), along with the known callyaerins A–G (6–12). The structures of the new compounds were unambiguously elucidated on the basis of one- and two-dimensional NMR spectroscopy and mass spectrometry. In addition, the structures of callyaerins D (9), F (11), and G (12), previously available in only small amounts, have been reinvestigated and revised. All compounds were tested in vitro against Mycobacterium tuberculosis, as well as against THP-1 (human acute monocytic leukemia) and MRC-5 (human fetal lung fibroblast) cell lines, in order to assess their general cytotoxicity. Callyaerins A (6) and B (7) showed potent anti-TB activity with MIC90 values of 2 and 5 μM, respectively. Callyaerin C (8) was found to be less active, with an MIC90 value of 40 μM. Callyaerin A (6), which showed the strongest anti-TB activity, was not cytotoxic to THP-1 or MRC-5 cells (IC50 > 10 μM), which highlights the potential of these compounds as promising anti-TB agents
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talaromins a and b new Cyclic Peptides from the endophytic fungus talaromyces wortmannii
Tetrahedron Letters, 2013Co-Authors: Robert Bara, Amal H. Aly, Victor Wray, Peter Proksch, Wenhan Lin, Abdessamad DebbabAbstract:Abstract Chemical investigation of the endophytic fungus Talaromyces wortmannii, isolated from Aloe vera, yielded two new Cyclic Peptides, talaromins A and B. Their structures were established on the basis of extensive NMR spectroscopic and mass spectrometric analysis. Both cycloPeptides contain ring systems comprised of six α-amino acid residues connected to a β-amino acid. The absolute configurations of the α-amino acids were determined by Marfey’s method.
Thomas Durek - One of the best experts on this subject based on the ideXlab platform.
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co expression of a cyclizing asparaginyl endopeptidase enables efficient production of Cyclic Peptides in planta
Journal of Experimental Botany, 2018Co-Authors: Simon Poon, David J. Craik, Thomas Durek, Karen S Harris, Edward K Gilding, Nicole L Van Der Weerden, Mark A Jackson, Owen C Mccorkelle, Marilyn A AndersonAbstract:Cyclotides are ultra-stable, backbone-cyclized plant defence Peptides that have attracted considerable interest in the pharmaceutical industry. This is due to their range of native bioactivities as well as their ability to stabilize other bioactive Peptides within their framework. However, a hindrance to their widespread application is the lack of scalable, cost-effective production strategies. Plant-based production is an attractive, benign option since all biosynthetic steps are performed in planta. Nonetheless, cyclization in non-cyclotide-producing plants is poor. Here, we show that Cyclic Peptides can be produced efficiently in Nicotiana benthamiana, one of the leading plant-based protein production platforms, by co-expressing cyclotide precursors with asparaginyl endopeptidases that catalyse peptide backbone cyclization. This approach was successful in a range of other plants (tobacco, bush bean, lettuce, and canola), either transiently or stably expressed, and was applicable to both native and engineered Cyclic Peptides. We also describe the use of the transgenic system to rapidly identify new asparaginyl endopeptidase cyclases and interrogate their substrate sequence requirements. Our results pave the way for exploiting cyclotides for pest protection in transgenic crops as well as large-scale production of Cyclic peptide pharmaceuticals in plants.
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Constrained Cyclic Peptides as Immunomodulatory Inhibitors of the CD2:CD58 Protein–Protein Interaction
2016Co-Authors: Rushikesh Sable, Thomas Durek, Veena Taneja, Sandeep Pallerla, David J. Craik, Ted Gauthier, Seetharama D. JoisAbstract:The interaction between the cell–cell adhesion proteins CD2 and CD58 plays a crucial role in lymphocyte recruitment to inflammatory sites, and inhibitors of this interaction have potential as immunomodulatory drugs in autoimmune diseases. Peptides from the CD2 adhesion domain were designed to inhibit CD2:CD58 interactions. To improve the stability of the Peptides, β-sheet epitopes from the CD2 region implicated in CD58 recognition were grafted into the Cyclic peptide frameworks of sunflower trypsin inhibitor and rhesus theta defensin. The designed multiCyclic Peptides were evaluated for their ability to modulate cell–cell interactions in three different cell adhesion assays, with one candidate, SFTI-a, showing potent activity in the nanomolar range (IC50: 51 nM). This peptide also suppresses the immune responses in T cells obtained from mice that exhibit the autoimmune disease rheumatoid arthritis. SFTI-a was resistant to thermal denaturation, as judged by circular dichroism spectroscopy and mass spectrometry, and had a half-life of ∼24 h in human serum. Binding of this peptide to CD58 was predicted by molecular docking studies and experimentally confirmed by surface plasmon resonance experiments. Our results suggest that Cyclic Peptides from natural sources are promising scaffolds for modulating protein–protein interactions that are typically difficult to target with small-molecule compounds
Victor Wray - One of the best experts on this subject based on the ideXlab platform.
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callyaerins from the marine sponge callyspongia aerizusa Cyclic Peptides with antitubercular activity
Journal of Natural Products, 2015Co-Authors: Georgios Daletos, Victor Wray, Wenhan Lin, Rainer Kalscheuer, Rudolf Hartmann, Nicole J. De Voogd, Hendrik Koliwerbrandl, Peter ProkschAbstract:Chemical investigation of the Indonesian sponge Callyspongia aerizusa afforded five new Cyclic Peptides, callyaerins I-M (1-5), along with the known callyaerins A-G (6-12). The structures of the new compounds were unambiguously elucidated on the basis of one- and two-dimensional NMR spectroscopy and mass spectrometry. In addition, the structures of callyaerins D (9), F (11), and G (12), previously available in only small amounts, have been reinvestigated and revised. All compounds were tested in vitro against Mycobacterium tuberculosis, as well as against THP-1 (human acute monocytic leukemia) and MRC-5 (human fetal lung fibroblast) cell lines, in order to assess their general cytotoxicity. Callyaerins A (6) and B (7) showed potent anti-TB activity with MIC₉₀ values of 2 and 5 μM, respectively. Callyaerin C (8) was found to be less active, with an MIC₉₀ value of 40 μM. Callyaerin A (6), which showed the strongest anti-TB activity, was not cytotoxic to THP-1 or MRC-5 cells (IC₅₀ > 10 μM), which highlights the potential of these compounds as promising anti-TB agents.
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Callyaerins from the Marine Sponge Callyspongia aerizusa: Cyclic Peptides with Antitubercular Activity
2015Co-Authors: Georgios Daletos, Victor Wray, Wenhan Lin, Rainer Kalscheuer, Hendrik Koliwer-brandl, Rudolf Hartmann, Nicole J. De Voogd, Peter ProkschAbstract:Chemical investigation of the Indonesian sponge Callyspongia aerizusa afforded five new Cyclic Peptides, callyaerins I–M (1–5), along with the known callyaerins A–G (6–12). The structures of the new compounds were unambiguously elucidated on the basis of one- and two-dimensional NMR spectroscopy and mass spectrometry. In addition, the structures of callyaerins D (9), F (11), and G (12), previously available in only small amounts, have been reinvestigated and revised. All compounds were tested in vitro against Mycobacterium tuberculosis, as well as against THP-1 (human acute monocytic leukemia) and MRC-5 (human fetal lung fibroblast) cell lines, in order to assess their general cytotoxicity. Callyaerins A (6) and B (7) showed potent anti-TB activity with MIC90 values of 2 and 5 μM, respectively. Callyaerin C (8) was found to be less active, with an MIC90 value of 40 μM. Callyaerin A (6), which showed the strongest anti-TB activity, was not cytotoxic to THP-1 or MRC-5 cells (IC50 > 10 μM), which highlights the potential of these compounds as promising anti-TB agents
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talaromins a and b new Cyclic Peptides from the endophytic fungus talaromyces wortmannii
Tetrahedron Letters, 2013Co-Authors: Robert Bara, Amal H. Aly, Victor Wray, Peter Proksch, Wenhan Lin, Abdessamad DebbabAbstract:Abstract Chemical investigation of the endophytic fungus Talaromyces wortmannii, isolated from Aloe vera, yielded two new Cyclic Peptides, talaromins A and B. Their structures were established on the basis of extensive NMR spectroscopic and mass spectrometric analysis. Both cycloPeptides contain ring systems comprised of six α-amino acid residues connected to a β-amino acid. The absolute configurations of the α-amino acids were determined by Marfey’s method.
Wenhan Lin - One of the best experts on this subject based on the ideXlab platform.
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callyaerins from the marine sponge callyspongia aerizusa Cyclic Peptides with antitubercular activity
Journal of Natural Products, 2015Co-Authors: Georgios Daletos, Victor Wray, Wenhan Lin, Rainer Kalscheuer, Rudolf Hartmann, Nicole J. De Voogd, Hendrik Koliwerbrandl, Peter ProkschAbstract:Chemical investigation of the Indonesian sponge Callyspongia aerizusa afforded five new Cyclic Peptides, callyaerins I-M (1-5), along with the known callyaerins A-G (6-12). The structures of the new compounds were unambiguously elucidated on the basis of one- and two-dimensional NMR spectroscopy and mass spectrometry. In addition, the structures of callyaerins D (9), F (11), and G (12), previously available in only small amounts, have been reinvestigated and revised. All compounds were tested in vitro against Mycobacterium tuberculosis, as well as against THP-1 (human acute monocytic leukemia) and MRC-5 (human fetal lung fibroblast) cell lines, in order to assess their general cytotoxicity. Callyaerins A (6) and B (7) showed potent anti-TB activity with MIC₉₀ values of 2 and 5 μM, respectively. Callyaerin C (8) was found to be less active, with an MIC₉₀ value of 40 μM. Callyaerin A (6), which showed the strongest anti-TB activity, was not cytotoxic to THP-1 or MRC-5 cells (IC₅₀ > 10 μM), which highlights the potential of these compounds as promising anti-TB agents.
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Callyaerins from the Marine Sponge Callyspongia aerizusa: Cyclic Peptides with Antitubercular Activity
2015Co-Authors: Georgios Daletos, Victor Wray, Wenhan Lin, Rainer Kalscheuer, Hendrik Koliwer-brandl, Rudolf Hartmann, Nicole J. De Voogd, Peter ProkschAbstract:Chemical investigation of the Indonesian sponge Callyspongia aerizusa afforded five new Cyclic Peptides, callyaerins I–M (1–5), along with the known callyaerins A–G (6–12). The structures of the new compounds were unambiguously elucidated on the basis of one- and two-dimensional NMR spectroscopy and mass spectrometry. In addition, the structures of callyaerins D (9), F (11), and G (12), previously available in only small amounts, have been reinvestigated and revised. All compounds were tested in vitro against Mycobacterium tuberculosis, as well as against THP-1 (human acute monocytic leukemia) and MRC-5 (human fetal lung fibroblast) cell lines, in order to assess their general cytotoxicity. Callyaerins A (6) and B (7) showed potent anti-TB activity with MIC90 values of 2 and 5 μM, respectively. Callyaerin C (8) was found to be less active, with an MIC90 value of 40 μM. Callyaerin A (6), which showed the strongest anti-TB activity, was not cytotoxic to THP-1 or MRC-5 cells (IC50 > 10 μM), which highlights the potential of these compounds as promising anti-TB agents
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talaromins a and b new Cyclic Peptides from the endophytic fungus talaromyces wortmannii
Tetrahedron Letters, 2013Co-Authors: Robert Bara, Amal H. Aly, Victor Wray, Peter Proksch, Wenhan Lin, Abdessamad DebbabAbstract:Abstract Chemical investigation of the endophytic fungus Talaromyces wortmannii, isolated from Aloe vera, yielded two new Cyclic Peptides, talaromins A and B. Their structures were established on the basis of extensive NMR spectroscopic and mass spectrometric analysis. Both cycloPeptides contain ring systems comprised of six α-amino acid residues connected to a β-amino acid. The absolute configurations of the α-amino acids were determined by Marfey’s method.
