Animals that specialise in blood feeding have particular challenges in obtaining their meal, wher... more Animals that specialise in blood feeding have particular challenges in obtaining their meal, whereby they impair blood hemostasis by promoting anticoagulation and vasodilation in order to facilitate feeding. These convergent selection pressures have been studied in a number of lineages, ranging from fleas to leeches. However, the vampire bat (Desmondus rotundus) is unstudied in regards to potential vasodilatory mechanisms of their feeding secretions (which are a type of venom). This is despite the intense investigations of their anticoagulant properties which have demonstrated that D. rotundus venom contains strong anticoagulant and proteolytic activities which delay the formation of blood clots and interfere with the blood coagulation cascade. In this study, we identified and tested a compound from D. rotundus venom that is similar in size and amino acid sequence to human calcitonin gene-related peptide (CGRP) which has potent vasodilatory properties. We found that the vampire bat-...
Cone snails are a diverse group of predatory marine invertebrates that deploy remarkably complex ... more Cone snails are a diverse group of predatory marine invertebrates that deploy remarkably complex venoms to rapidly paralyse worm, mollusc or fish prey. ω-Conotoxins are neurotoxic peptides from cone snail venoms that inhibit Ca2.2 voltage-gated calcium channel, demonstrating potential for pain management via intrathecal (IT) administration. Here, we isolated and characterized two novel ω-conotoxins, MoVIA and MoVIB from Conus moncuri, the first to be identified in vermivorous (worm-hunting) cone snails. MoVIA and MoVIB potently inhibited human Ca2.2 in fluorimetric assays and rat Ca2.2 in patch clamp studies, and both potently displaced radiolabeled ω-conotoxin GVIA (I-GVIA) from human SH-SY5Y cells and fish brain membranes (IC 2-9 pM). Intriguingly, an arginine at position 13 in MoVIA and MoVIB replaced the functionally critical tyrosine found in piscivorous ω-conotoxins. To investigate its role, we synthesized MoVIB-[R13Y] and MVIIA-[Y13R]. Interestingly, MVIIA-[Y13R] completely l...
The Tasmanian devil faces extinction due to devil facial tumour disease (DFTD), a highly transmit... more The Tasmanian devil faces extinction due to devil facial tumour disease (DFTD), a highly transmittable clonal form of cancer without available treatment. In this study, we report the cell-autonomous antiproliferative and cytotoxic activities exhibited by the spider peptide gomesin (AgGom) and gomesin-like homologue (HiGom) in DFTD cells. Mechanistically, both peptides caused a significant reduction at G0/G1 phase, in correlation with an augmented expression of the cell cycle inhibitory proteins p53, p27, p21, necrosis, exacerbated generation of reactive oxygen species and diminished mitochondrial membrane potential, all hallmarks of cellular stress. The screening of a novel panel of AgGom-analogues revealed that, unlike changes in the hydrophobicity and electrostatic surface, the cytotoxic potential of the gomesin analogues in DFTD cells lies on specific arginine substitutions in the eight and nine positions and alanine replacement in three, five and 12 positions. In conclusion, the...
Peptide dendrimers are a novel class of precisely defined macromolecules of emerging interest. He... more Peptide dendrimers are a novel class of precisely defined macromolecules of emerging interest. Here, we describe the synthesis, structure, binding affinity, receptor selectivity, functional activity, and antinociceptive properties of oxytocin-related dendrimers containing up to 16 copies of [Lys8]-oxytocin or LVT. These were generated using a copper(i)-catalyzed azide–alkyne cycloaddition (CuAAc) reaction with azido-pegylated LVT peptides on an alkyne–polylysine scaffold. 2D NMR analysis demonstrated that each attached LVT ligand was freely rotating and maintained identical 3D structures in each dendrimeric macromolecule. The binding affinity Ki at the oxytocin receptor increased approximately 17-, 12-, 3-, and 1.5-fold respectively for the 2-, 4-, 8-, and 16-mer dendrimeric LVT conjugates, compared with monomer azido-pegylated LVT (Ki = 9.5 nM), consistent with a multivalency effect. A similar trend in affinity was also observed at the related human V1a, V1b, and V2 receptors, with...
Journal of peptide science : an official publication of the European Peptide Society, Jan 22, 2016
Peptide dendrimers are a novel class of macromolecules of emerging interest with the potential of... more Peptide dendrimers are a novel class of macromolecules of emerging interest with the potential of delayed renal clearance due to their molecular size and enhanced activity due to the multivalency effect. In this work, an active analogue of the disulfide-rich χ-conotoxin χ-MrIA (χ-MrIA), a norepinephrine reuptake (norepinephrine transporter) inhibitor, was grafted onto a polylysine dendron. Dendron decoration was achieved by employing copper-catalyzed alkyne-azide cycloaddition with azido-PEG chain-modified χ-MrIA analogues, leading to homogenous 4-mer and 8-mer χ-MrIA dendrimers with molecular weights ranging from 8 to 22 kDa. These dendrimers were investigated for their impact on peptide secondary structure, in vitro functional activity, and potential anti-allodynia in vivo. NMR studies showed that the χ-MrIA tertiary structure was maintained in the χ-MrIA dendrimers. In a functional norepinephrine transporter reuptake assay, χ-MrIA dendrimers showed slightly increased potency rela...
Natriuretic peptides (NP) play important roles in human cardiac physiology through their guanylyl... more Natriuretic peptides (NP) play important roles in human cardiac physiology through their guanylyl cyclase receptors NPR-A and NPR-B. Described herein is a bifunctional O-glycosylated natriuretic peptide, TcNPa, from Tropidechis carinatus venom and it unusually targets both NPR-A and NPR-B. Characterization using specific glycosidases and ETD-MS identified the glycan as galactosyl-β(1-3)-N-acetylgalactosamine (Gal-GalNAc) and was α-linked to the C-terminal threonine residue. TcNPa contains the characteristic NP 17-membered disulfide ring with conserved phenylalanine and arginine residues. Both glycosylated and nonglycosylated forms were synthesized by Fmoc solid-phase peptide synthesis and NMR analysis identified an α-helix within the disulfide ring containing the putative pharmacophore for NPR-A. Surprisingly, both forms activated NPR-A and NPR-B and were relatively resistant towards proteolytic degradation in plasma. This work will underpin the future development of bifunctional NP peptide mimetics.
with specic help available everywhere you see the i O symbol. The following versions of software ... more with specic help available everywhere you see the i O symbol. The following versions of software and data (see references i O) were used in the production of this report:
The design of disulfide bond mimetics is an important strategy for optimising cysteine‐rich pepti... more The design of disulfide bond mimetics is an important strategy for optimising cysteine‐rich peptides in drug development. Mimetics of the drug lead conotoxin MrIA, in which one disulfide bond is selectively replaced of by a 1,4‐disubstituted‐1,2,3‐triazole bridge, are described. Sequential copper‐catalyzed azide–alkyne cycloaddition (CuAAC; click reaction) followed by disulfide formation resulted in the regioselective syntheses of triazole–disulfide hybrid MrIA analogues. Mimetics with a triazole replacing the Cys4–Cys13 disulfide bond retained tertiary structure and full in vitro and in vivo activity as norepinephrine reuptake inhibitors. Importantly, these mimetics are resistant to reduction in the presence of glutathione, thus resulting in improved plasma stability and increased suitability for drug development.
Die Synthese von Disulfidbrückenmimetika ist eine wichtige Strategie zur Optimierung disulfidreic... more Die Synthese von Disulfidbrückenmimetika ist eine wichtige Strategie zur Optimierung disulfidreicher Peptide beim Wirkstoff-Design. Wir beschreiben Mimetika des Conotoxins MrIA, die durch selektiven Austausch einzelner Disulfidbindungen gegen Brücken aus einem 1,4-disubstituierten 1,2,3-Triazol erhalten wurden. Eine sequenzielle, Kupfer-katalysierte Azid-Alkin-Klick-Reaktion (CuAAC) mit nachfolgender Disulfidbildung führte regioselektiv zu hybriden Triazol-Disulfid-Analoga von MrIA. Mimetika, in denen Triazol die Cys4-Cys13-Disulfidbindung ersetzte, behielten ihre Tertiärstruktur sowie die volle In-vitro-und In-vivo-Aktivität als Inhibitoren der Noradrenalin-Wiederaufnahme bei. Diese Mimetika sind resistent gegen Reduktion mit Glutathion, was zu einer verbesserten Plasmastabilität führt und dadurch interessant für die Wirkstoff-Entwicklung ist. Schema 1. A) Disulfid-Faltung führt zu mehreren Isomeren, disulfidreiche Peptide sind anfällig für "S-S-Austausch". B) Disulfid-/1,2,3-Triazol-Faltung liefert nur eines der mçglichen Isomere.
We suggest that subscribers photocopy these corrections and insert the photocopies at the appropr... more We suggest that subscribers photocopy these corrections and insert the photocopies at the appropriate places where the article to be corrected originally appeared. Authors are urged to introduce these corrections into any reprints they distribute. Secondary (abstract) services are urged to carry notice of these corrections as prominently as they carried the original abstracts.
Animals that specialise in blood feeding have particular challenges in obtaining their meal, wher... more Animals that specialise in blood feeding have particular challenges in obtaining their meal, whereby they impair blood hemostasis by promoting anticoagulation and vasodilation in order to facilitate feeding. These convergent selection pressures have been studied in a number of lineages, ranging from fleas to leeches. However, the vampire bat (Desmondus rotundus) is unstudied in regards to potential vasodilatory mechanisms of their feeding secretions (which are a type of venom). This is despite the intense investigations of their anticoagulant properties which have demonstrated that D. rotundus venom contains strong anticoagulant and proteolytic activities which delay the formation of blood clots and interfere with the blood coagulation cascade. In this study, we identified and tested a compound from D. rotundus venom that is similar in size and amino acid sequence to human calcitonin gene-related peptide (CGRP) which has potent vasodilatory properties. We found that the vampire bat-...
Cone snails are a diverse group of predatory marine invertebrates that deploy remarkably complex ... more Cone snails are a diverse group of predatory marine invertebrates that deploy remarkably complex venoms to rapidly paralyse worm, mollusc or fish prey. ω-Conotoxins are neurotoxic peptides from cone snail venoms that inhibit Ca2.2 voltage-gated calcium channel, demonstrating potential for pain management via intrathecal (IT) administration. Here, we isolated and characterized two novel ω-conotoxins, MoVIA and MoVIB from Conus moncuri, the first to be identified in vermivorous (worm-hunting) cone snails. MoVIA and MoVIB potently inhibited human Ca2.2 in fluorimetric assays and rat Ca2.2 in patch clamp studies, and both potently displaced radiolabeled ω-conotoxin GVIA (I-GVIA) from human SH-SY5Y cells and fish brain membranes (IC 2-9 pM). Intriguingly, an arginine at position 13 in MoVIA and MoVIB replaced the functionally critical tyrosine found in piscivorous ω-conotoxins. To investigate its role, we synthesized MoVIB-[R13Y] and MVIIA-[Y13R]. Interestingly, MVIIA-[Y13R] completely l...
The Tasmanian devil faces extinction due to devil facial tumour disease (DFTD), a highly transmit... more The Tasmanian devil faces extinction due to devil facial tumour disease (DFTD), a highly transmittable clonal form of cancer without available treatment. In this study, we report the cell-autonomous antiproliferative and cytotoxic activities exhibited by the spider peptide gomesin (AgGom) and gomesin-like homologue (HiGom) in DFTD cells. Mechanistically, both peptides caused a significant reduction at G0/G1 phase, in correlation with an augmented expression of the cell cycle inhibitory proteins p53, p27, p21, necrosis, exacerbated generation of reactive oxygen species and diminished mitochondrial membrane potential, all hallmarks of cellular stress. The screening of a novel panel of AgGom-analogues revealed that, unlike changes in the hydrophobicity and electrostatic surface, the cytotoxic potential of the gomesin analogues in DFTD cells lies on specific arginine substitutions in the eight and nine positions and alanine replacement in three, five and 12 positions. In conclusion, the...
Peptide dendrimers are a novel class of precisely defined macromolecules of emerging interest. He... more Peptide dendrimers are a novel class of precisely defined macromolecules of emerging interest. Here, we describe the synthesis, structure, binding affinity, receptor selectivity, functional activity, and antinociceptive properties of oxytocin-related dendrimers containing up to 16 copies of [Lys8]-oxytocin or LVT. These were generated using a copper(i)-catalyzed azide–alkyne cycloaddition (CuAAc) reaction with azido-pegylated LVT peptides on an alkyne–polylysine scaffold. 2D NMR analysis demonstrated that each attached LVT ligand was freely rotating and maintained identical 3D structures in each dendrimeric macromolecule. The binding affinity Ki at the oxytocin receptor increased approximately 17-, 12-, 3-, and 1.5-fold respectively for the 2-, 4-, 8-, and 16-mer dendrimeric LVT conjugates, compared with monomer azido-pegylated LVT (Ki = 9.5 nM), consistent with a multivalency effect. A similar trend in affinity was also observed at the related human V1a, V1b, and V2 receptors, with...
Journal of peptide science : an official publication of the European Peptide Society, Jan 22, 2016
Peptide dendrimers are a novel class of macromolecules of emerging interest with the potential of... more Peptide dendrimers are a novel class of macromolecules of emerging interest with the potential of delayed renal clearance due to their molecular size and enhanced activity due to the multivalency effect. In this work, an active analogue of the disulfide-rich χ-conotoxin χ-MrIA (χ-MrIA), a norepinephrine reuptake (norepinephrine transporter) inhibitor, was grafted onto a polylysine dendron. Dendron decoration was achieved by employing copper-catalyzed alkyne-azide cycloaddition with azido-PEG chain-modified χ-MrIA analogues, leading to homogenous 4-mer and 8-mer χ-MrIA dendrimers with molecular weights ranging from 8 to 22 kDa. These dendrimers were investigated for their impact on peptide secondary structure, in vitro functional activity, and potential anti-allodynia in vivo. NMR studies showed that the χ-MrIA tertiary structure was maintained in the χ-MrIA dendrimers. In a functional norepinephrine transporter reuptake assay, χ-MrIA dendrimers showed slightly increased potency rela...
Natriuretic peptides (NP) play important roles in human cardiac physiology through their guanylyl... more Natriuretic peptides (NP) play important roles in human cardiac physiology through their guanylyl cyclase receptors NPR-A and NPR-B. Described herein is a bifunctional O-glycosylated natriuretic peptide, TcNPa, from Tropidechis carinatus venom and it unusually targets both NPR-A and NPR-B. Characterization using specific glycosidases and ETD-MS identified the glycan as galactosyl-β(1-3)-N-acetylgalactosamine (Gal-GalNAc) and was α-linked to the C-terminal threonine residue. TcNPa contains the characteristic NP 17-membered disulfide ring with conserved phenylalanine and arginine residues. Both glycosylated and nonglycosylated forms were synthesized by Fmoc solid-phase peptide synthesis and NMR analysis identified an α-helix within the disulfide ring containing the putative pharmacophore for NPR-A. Surprisingly, both forms activated NPR-A and NPR-B and were relatively resistant towards proteolytic degradation in plasma. This work will underpin the future development of bifunctional NP peptide mimetics.
with specic help available everywhere you see the i O symbol. The following versions of software ... more with specic help available everywhere you see the i O symbol. The following versions of software and data (see references i O) were used in the production of this report:
The design of disulfide bond mimetics is an important strategy for optimising cysteine‐rich pepti... more The design of disulfide bond mimetics is an important strategy for optimising cysteine‐rich peptides in drug development. Mimetics of the drug lead conotoxin MrIA, in which one disulfide bond is selectively replaced of by a 1,4‐disubstituted‐1,2,3‐triazole bridge, are described. Sequential copper‐catalyzed azide–alkyne cycloaddition (CuAAC; click reaction) followed by disulfide formation resulted in the regioselective syntheses of triazole–disulfide hybrid MrIA analogues. Mimetics with a triazole replacing the Cys4–Cys13 disulfide bond retained tertiary structure and full in vitro and in vivo activity as norepinephrine reuptake inhibitors. Importantly, these mimetics are resistant to reduction in the presence of glutathione, thus resulting in improved plasma stability and increased suitability for drug development.
Die Synthese von Disulfidbrückenmimetika ist eine wichtige Strategie zur Optimierung disulfidreic... more Die Synthese von Disulfidbrückenmimetika ist eine wichtige Strategie zur Optimierung disulfidreicher Peptide beim Wirkstoff-Design. Wir beschreiben Mimetika des Conotoxins MrIA, die durch selektiven Austausch einzelner Disulfidbindungen gegen Brücken aus einem 1,4-disubstituierten 1,2,3-Triazol erhalten wurden. Eine sequenzielle, Kupfer-katalysierte Azid-Alkin-Klick-Reaktion (CuAAC) mit nachfolgender Disulfidbildung führte regioselektiv zu hybriden Triazol-Disulfid-Analoga von MrIA. Mimetika, in denen Triazol die Cys4-Cys13-Disulfidbindung ersetzte, behielten ihre Tertiärstruktur sowie die volle In-vitro-und In-vivo-Aktivität als Inhibitoren der Noradrenalin-Wiederaufnahme bei. Diese Mimetika sind resistent gegen Reduktion mit Glutathion, was zu einer verbesserten Plasmastabilität führt und dadurch interessant für die Wirkstoff-Entwicklung ist. Schema 1. A) Disulfid-Faltung führt zu mehreren Isomeren, disulfidreiche Peptide sind anfällig für "S-S-Austausch". B) Disulfid-/1,2,3-Triazol-Faltung liefert nur eines der mçglichen Isomere.
We suggest that subscribers photocopy these corrections and insert the photocopies at the appropr... more We suggest that subscribers photocopy these corrections and insert the photocopies at the appropriate places where the article to be corrected originally appeared. Authors are urged to introduce these corrections into any reprints they distribute. Secondary (abstract) services are urged to carry notice of these corrections as prominently as they carried the original abstracts.
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