The Experts below are selected from a list of 7026 Experts worldwide ranked by ideXlab platform
David J. Adams - One of the best experts on this subject based on the ideXlab platform.
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dimerization of α Conotoxins as a strategy to enhance the inhibition of the human α7 and α9α10 nicotinic acetylcholine receptors
Journal of Medicinal Chemistry, 2020Co-Authors: Jiazhen Liang, Hanshen Tae, Tao Jiang, David J. AdamsAbstract:The affinity of α-Conotoxins, a class of nicotinic acetylcholine receptor (nAChR) peptide inhibitors, can be enhanced by dendrimerization. It has been hypothesized that this improvement arose from simultaneous binding of the α-Conotoxins to several spatially adjacent sites. We here engineered several α-Conotoxin dimers using a linker length compatible between neighboring binding sites on the same receptor. Remarkably, the dimer of α-Conotoxin PeIA compared to the monomer displayed an increase in potency by 11-fold (IC50 = 1.9 nM) for the human α9α10 nAChR. The dimerization of α-Conotoxin RgIA# resulted in a dual inhibitor that targets both α9α10 and α7 nAChR subtypes with an IC50 = ∼50 nM. The RgIA# dimer is therapeutically interesting because it is the first dual inhibitor that potently and selectively inhibits these two nAChR subtypes, which are both involved in the etiology of several cancers. We propose that the dimerization of α-Conotoxins is a simpler and efficient alternative strategy to dendrimers for enhancing the activity of α-Conotoxins.
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key structural determinants in the agonist binding loops of human β2 and β4 nicotinic acetylcholine receptor subunits contribute to α3β4 subtype selectivity of α Conotoxins
Journal of Biological Chemistry, 2016Co-Authors: David J. Adams, Hanshen Tae, Hartmut Cuny, Shiva N KompellaAbstract:α-Conotoxins represent a large group of pharmacologically active peptides that antagonize nicotinic acetylcholine receptors (nAChRs). The α3β4 nAChR, a predominant subtype in the peripheral nervous system, has been implicated in various pathophysiological conditions. As many α-Conotoxins have multiple pharmacological targets, compounds specifically targeting individual nAChR subtypes are needed. In this study, we performed mutational analyses to evaluate the key structural components of human β2 and β4 nAChR subunits that determine α-Conotoxin selectivity for α3β4 nAChR. α-Conotoxin RegIIA was used to evaluate the impact of non-conserved human β2 and β4 residues on peptide affinity. Two mutations, α3β2[T59K] and α3β2[S113R], strongly enhanced RegIIA affinity compared with wild-type α3β2, as seen by substantially increased inhibitory potency and slower off-rate kinetics. Opposite point mutations in α3β4 had the contrary effect, emphasizing the importance of loop D residue 59 and loop E residue 113 as determinants for RegIIA affinity. Molecular dynamics simulation revealed the side chains of β4 Lys59 and β4 Arg113 formed hydrogen bonds with RegIIA loop 2 atoms, whereas the β2 Thr59 and β2 Ser113 side chains were not long enough to form such interactions. Residue β4 Arg113 has been identified for the first time as a crucial component facilitating antagonist binding. Another α-Conotoxin, AuIB, exhibited low activity at human α3β2 and α3β4 nAChRs. Molecular dynamics simulation indicated the key interactions with the β subunit are different to RegIIA. Taken together, these data elucidate the interactions with specific individual β subunit residues that critically determine affinity and pharmacological activity of α-Conotoxins RegIIA and AuIB at human nAChRs.
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Conotoxins targeting neuronal voltage gated sodium channel subtypes potential analgesics
Toxins, 2012Co-Authors: Oliver Knapp, Jeffrey R Mcarthur, David J. AdamsAbstract:Voltage-gated sodium channels (VGSC) are the primary mediators of electrical signal amplification and propagation in excitable cells. VGSC subtypes are diverse, with different biophysical and pharmacological properties, and varied tissue distribution. Altered VGSC expression and/or increased VGSC activity in sensory neurons is characteristic of inflammatory and neuropathic pain states. Therefore, VGSC modulators could be used in prospective analgesic compounds. VGSCs have specific binding sites for four Conotoxin families: μ-, μO-, δ- and ί-Conotoxins. Various studies have identified that the binding site of these peptide toxins is restricted to well-defined areas or domains. To date, only the μ- and μO-family exhibit analgesic properties in animal pain models. This review will focus on Conotoxins from the μ- and μO-families that act on neuronal VGSCs. Examples of how these Conotoxins target various pharmacologically important neuronal ion channels, as well as potential problems with the development of drugs from Conotoxins, will be discussed.
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isolation characterization and total regioselective synthesis of the novel μo Conotoxin mfvia from conus magnificus that targets voltage gated sodium channels
Biochemical Pharmacology, 2012Co-Authors: Irina Vetter, Paul Francis Alewood, Zoltan Dekan, David J. Adams, Oliver Knapp, Richard J. LewisAbstract:The μO-Conotoxins are notable for their unique selectivity for Na(v)1.8 over other sodium channel isoforms, making them attractive drug leads for the treatment of neuropathic pain. We describe the discovery of a novel μO-Conotoxin, MfVIA, from the venom of Conus magnificus using high-throughput screening approaches. MfVIA was found to be a hydrophobic 32-residue peptide (amino acid sequence RDCQEKWEYCIVPILGFVYCCPGLICGPFVCV) with highest sequence homology to μO-Conotoxin MrVIB. To overcome the synthetic challenges posed by μO-Conotoxins due to their hydrophobic nature and difficult folding, we developed a novel regioselective approach for the synthesis of μO-Conotoxins. Performing selective oxidative deprotections of the cysteine side-chain protecting groups of the fully protected peptide allowed manipulations in organic solvents with no chromatography required between steps. Using this approach, we obtained correctly folded MfVIA with increased synthetic yields. Biological activity of MfVIA was assessed using membrane potential-sensitive dyes and electrophysiological recording techniques. MfVIA preferentially inhibits Na(v)1.8 (IC₅₀ 95.9±74.3 nM) and Na(v)1.4 (IC₅₀ 81±16 nM), with significantly lower affinity for other Na(v) subtypes (IC₅₀ 431-6203 nM; Na(v)1.5>1.6∼1.7∼1.3∼1.1∼1.2). This improved approach to μO-Conotoxin synthesis will facilitate the optimization of μO-Conotoxins as novel analgesic molecules to improve pain management.
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analgesic Conotoxins block and g protein coupled receptor modulation of n type cav2 2 calcium channels
British Journal of Pharmacology, 2012Co-Authors: David J. Adams, Brid P Callaghan, Geza BereckiAbstract:Conotoxins (conopeptides) are small disulfide bonded peptides from the venom of marine cone snails. These peptides target a wide variety of membrane receptors, ion channels and transporters, and have enormous potential for a range of pharmaceutical applications. Structurally related ω-Conotoxins bind directly to and selectively inhibit neuronal (N)-type voltage-gated calcium channels (VGCCs) of nociceptive primary afferent neurones. Among these, ω-Conotoxin MVIIA (Prialt) is approved by the Food and Drug Administration (FDA) as an alternative intrathecal analgesic for the management of chronic intractable pain, particularly in patients refractory to opioids. A series of newly discovered ω-Conotoxins from Conus catus, including CVID–F, are potent and selective antagonists of N-type VGCCs. In spinal cord slices, these peptides reversibly inhibit excitatory synaptic transmission between primary afferents and dorsal horn superficial lamina neurones, and in the rat partial sciatic nerve ligation model of neuropathic pain, significantly reduce allodynic behaviour. Another family of Conotoxins, the α-Conotoxins, are competitive antagonists of mammalian nicotinic acetylcholine receptors (nAChRs). α-Conotoxins Vc1.1 and RgIA possess two disulfide bonds and are currently in development as a treatment for neuropathic pain. It was initially proposed that the primary target of these peptides is the α9α10 neuronal nAChR. Surprisingly, however, α-Conotoxins Vc1.1, RgIA and PeIA more potently inhibit N-type VGCC currents via a GABAB GPCR mechanism in rat sensory neurones. This inhibition is largely voltage-independent and involves complex intracellular signalling. Understanding the molecular mechanisms of Conotoxin action will lead to new ways to regulate VGCC block and modulation in normal and diseased states of the nervous system.
Baldomero M. Olivera - One of the best experts on this subject based on the ideXlab platform.
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curses or cures a review of the numerous benefits versus the biosecurity concerns of Conotoxin research
Biomedicines, 2020Co-Authors: Walden E Bjornyoshimoto, Baldomero M. Olivera, Mark Yandell, Lars Ellgaard, Helena Safavihemami, Michael J Mcintosh, Iris Bea L RamiroAbstract:Conotoxins form a diverse group of peptide toxins found in the venom of predatory marine cone snails. Decades of Conotoxin research have provided numerous measurable scientific and societal benefits. These include their use as a drug, diagnostic agent, drug leads, and research tools in neuroscience, pharmacology, biochemistry, structural biology, and molecular evolution. Human envenomations by cone snails are rare but can be fatal. Death by envenomation is likely caused by a small set of toxins that induce muscle paralysis of the diaphragm, resulting in respiratory arrest. The potency of these toxins led to concerns regarding the potential development and use of Conotoxins as biological weapons. To address this, various regulatory measures have been introduced that limit the use and access of Conotoxins within the research community. Some of these regulations apply to all of the ≈200,000 Conotoxins predicted to exist in nature of which less than 0.05% are estimated to have any significant toxicity in humans. In this review we provide an overview of the many benefits of Conotoxin research, and contrast these to the perceived biosecurity concerns of Conotoxins and research thereof.
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ero1 mediated reoxidation of protein disulfide isomerase accelerates the folding of cone snail toxins
International Journal of Molecular Sciences, 2018Co-Authors: Henrik Obrien, Baldomero M. Olivera, Shingo Kanemura, Masaki Okumura, Robert P Baskin, Pradip K Bandyopadhyay, Lars Ellgaard, Kenji Inaba, Helena SafavihemamiAbstract:Disulfide-rich peptides are highly abundant in nature and their study has provided fascinating insight into protein folding, structure and function. Venomous cone snails belong to a group of organisms that express one of the largest sets of disulfide-rich peptides (Conotoxins) found in nature. The diversity of structural scaffolds found for Conotoxins suggests that specialized molecular adaptations have evolved to ensure their efficient folding and secretion. We recently showed that canonical protein disulfide isomerase (PDI) and a Conotoxin-specific PDI (csPDI) are ubiquitously expressed in the venom gland of cone snails and play a major role in Conotoxin folding. Here, we identify cone snail endoplasmic reticulum oxidoreductin-1 (Conus Ero1) and investigate its role in the oxidative folding of Conotoxins through reoxidation of cone snail PDI and csPDI. We show that Conus Ero1 preferentially reoxidizes PDI over csPDI, suggesting that the reoxidation of csPDI may rely on an Ero1-independent molecular pathway. Despite the preferential reoxidation of PDI over csPDI, the combinatorial effect of Ero1 and csPDI provides higher folding yields than Ero1 and PDI. We further demonstrate that the highest in vitro folding rates of two model Conotoxins are achieved when all three enzymes are present, indicating that these enzymes may act synergistically. Our findings provide new insight into the generation of one of the most diverse classes of disulfide-rich peptides and may improve current in vitro approaches for the production of venom peptides for pharmacological studies.
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The Venom Repertoire of Conus gloriamaris (Chemnitz, 1777), the Glory of the Sea
MDPI AG, 2017Co-Authors: Samuel D. Robinson, Baldomero M. Olivera, Mark Yandell, Pradip K Bandyopadhyay, Helena Safavi-hemamiAbstract:The marine cone snail Conus gloriamaris is an iconic species. For over two centuries, its shell was one of the most prized and valuable natural history objects in the world. Today, cone snails have attracted attention for their remarkable venom components. Many Conotoxins are proving valuable as research tools, drug leads, and drugs. In this article, we present the venom gland transcriptome of C. gloriamaris, revealing this species’ Conotoxin repertoire. More than 100 Conotoxin sequences were identified, representing a valuable resource for future drug discovery efforts
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the tetrodotoxin receptor of voltage gated sodium channels perspectives from interactions with μ Conotoxins
Marine Drugs, 2010Co-Authors: Robert J French, Doju Yoshikami, Michael F Sheets, Baldomero M. OliveraAbstract:Neurotoxin receptor site 1, in the outer vestibule of the conducting pore of voltage-gated sodium channels (VGSCs), was first functionally defined by its ability to bind the guanidinium-containing agents, tetrodotoxin (TTX) and saxitoxin (STX). Subsequent studies showed that peptide μ-Conotoxins competed for binding at site 1. All of these natural inhibitors block single sodium channels in an all-or-none manner on binding. With the discovery of an increasing variety of μ-Conotoxins, and the synthesis of numerous derivatives, observed interactions between the channel and these different ligands have become more complex. Certain μ-Conotoxin derivatives block single-channel currents partially, rather than completely, thus enabling the demonstration of interactions between the bound toxin and the channel’s voltage sensor. Most recently, the relatively small μ-Conotoxin KIIIA (16 amino acids) and its variants have been shown to bind simultaneously with TTX and exhibit both synergistic and antagonistic interactions with TTX. These interactions raise new pharmacological possibilities and place new constraints on the possible structures of the bound complexes of VGSCs with these toxins.
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nmr structure determination of α Conotoxin buia a novel neuronal nicotinic acetylcholine receptor antagonist with an unusual 4 4 disulfide scaffold
Biochemical and Biophysical Research Communications, 2006Co-Authors: Seung Wook Chi, Baldomero M. Olivera, Do Hyoung Kim, Michael J Mcintosh, Kyouhoon HanAbstract:We have determined a high-resolution three-dimensional structure of alpha-Conotoxin BuIA, a 13-residue peptide toxin isolated from Conus bullatus. Despite its unusual 4/4 disulfide bond layout alpha-Conotoxin BuIA exhibits strong antagonistic activity at alpha6/alpha3beta2beta3, alpha3beta2, and alpha3beta4 nAChR subtypes like some alpha4/7 Conotoxins. alpha-Conotoxin BuIA lacks the C-terminal beta-turn present within the second disulfide loop of alpha4/7 Conotoxins, having only a "pseudo omega-shaped" molecular topology. Nevertheless, it contains a functionally critical two-turn helix motif, a feature ubiquitously found in alpha4/7 Conotoxins. Such an aspect seems mainly responsible for similarities in the receptor recognition profile of alpha-Conotoxin BuIA to alpha4/7 Conotoxins. Structural comparison of alpha-Conotoxin BuIA with alpha4/7 Conotoxins and alpha4/3 Conotoxin ImI suggests that presence of the second helical turn portion of the two-turn helix motif in alpha4/7 and alpha4/4 Conotoxins may be important for binding to the alpha3 and/or alpha6 subunit of nAChR.
Michael J Mcintosh - One of the best experts on this subject based on the ideXlab platform.
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curses or cures a review of the numerous benefits versus the biosecurity concerns of Conotoxin research
Biomedicines, 2020Co-Authors: Walden E Bjornyoshimoto, Baldomero M. Olivera, Mark Yandell, Lars Ellgaard, Helena Safavihemami, Michael J Mcintosh, Iris Bea L RamiroAbstract:Conotoxins form a diverse group of peptide toxins found in the venom of predatory marine cone snails. Decades of Conotoxin research have provided numerous measurable scientific and societal benefits. These include their use as a drug, diagnostic agent, drug leads, and research tools in neuroscience, pharmacology, biochemistry, structural biology, and molecular evolution. Human envenomations by cone snails are rare but can be fatal. Death by envenomation is likely caused by a small set of toxins that induce muscle paralysis of the diaphragm, resulting in respiratory arrest. The potency of these toxins led to concerns regarding the potential development and use of Conotoxins as biological weapons. To address this, various regulatory measures have been introduced that limit the use and access of Conotoxins within the research community. Some of these regulations apply to all of the ≈200,000 Conotoxins predicted to exist in nature of which less than 0.05% are estimated to have any significant toxicity in humans. In this review we provide an overview of the many benefits of Conotoxin research, and contrast these to the perceived biosecurity concerns of Conotoxins and research thereof.
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a novel inhibitor of α9α10 nicotinic acetylcholine receptors from conus vexillum delineates a new Conotoxin superfamily
PLOS ONE, 2013Co-Authors: Sulan Luo, Raymond S. Norton, Dongting Zhangsun, Xiaopeng Zhu, Sean Christensen, Sandeep Chhabra, Michael J McintoshAbstract:Conotoxins (CTxs) selectively target a range of ion channels and receptors, making them widely used tools for probing nervous system function. Conotoxins have been previously grouped into superfamilies according to signal sequence and into families based on their cysteine framework and biological target. Here we describe the cloning and characterization of a new Conotoxin, from Conus vexillum, named αB-Conotoxin VxXXIVA. The peptide does not belong to any previously described Conotoxin superfamily and its arrangement of Cys residues is unique among conopeptides. Moreover, in contrast to previously characterized conopeptide toxins, which are expressed initially as prepropeptide precursors with a signal sequence, a ‘‘pro’’ region, and the toxin-encoding region, the precursor sequence of αB-VxXXIVA lacks a ‘‘pro’’ region. The predicted 40-residue mature peptide, which contains four Cys, was synthesized in each of the three possible disulfide arrangements. Investigation of the mechanism of action of αB-VxXXIVA revealed that the peptide is a nicotinic acetylcholine receptor (nAChR) antagonist with greatest potency against the α9α10 subtype. 1H nuclear magnetic resonance (NMR) spectra indicated that all three αB-VxXXIVA isomers were poorly structured in aqueous solution. This was consistent with circular dichroism (CD) results which showed that the peptides were unstructured in buffer, but adopted partially helical conformations in aqueous trifluoroethanol (TFE) solution. The α9α10 nAChR is an important target for the development of analgesics and cancer chemotherapeutics, and αB-VxXXIVA represents a novel ligand with which to probe the structure and function of this protein.
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nmr structure determination of α Conotoxin buia a novel neuronal nicotinic acetylcholine receptor antagonist with an unusual 4 4 disulfide scaffold
Biochemical and Biophysical Research Communications, 2006Co-Authors: Seung Wook Chi, Baldomero M. Olivera, Do Hyoung Kim, Michael J Mcintosh, Kyouhoon HanAbstract:We have determined a high-resolution three-dimensional structure of alpha-Conotoxin BuIA, a 13-residue peptide toxin isolated from Conus bullatus. Despite its unusual 4/4 disulfide bond layout alpha-Conotoxin BuIA exhibits strong antagonistic activity at alpha6/alpha3beta2beta3, alpha3beta2, and alpha3beta4 nAChR subtypes like some alpha4/7 Conotoxins. alpha-Conotoxin BuIA lacks the C-terminal beta-turn present within the second disulfide loop of alpha4/7 Conotoxins, having only a "pseudo omega-shaped" molecular topology. Nevertheless, it contains a functionally critical two-turn helix motif, a feature ubiquitously found in alpha4/7 Conotoxins. Such an aspect seems mainly responsible for similarities in the receptor recognition profile of alpha-Conotoxin BuIA to alpha4/7 Conotoxins. Structural comparison of alpha-Conotoxin BuIA with alpha4/7 Conotoxins and alpha4/3 Conotoxin ImI suggests that presence of the second helical turn portion of the two-turn helix motif in alpha4/7 and alpha4/4 Conotoxins may be important for binding to the alpha3 and/or alpha6 subunit of nAChR.
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solution conformation of a neuronal nicotinic acetylcholine receptor antagonist α Conotoxin omia that discriminates α3 vs α6 nachr subtypes
Biochemical and Biophysical Research Communications, 2006Co-Authors: Seung Wook Chi, Baldomero M. Olivera, Do Hyoung Kim, Michael J Mcintosh, Kyouhoon HanAbstract:{alpha}-Conotoxin OmIA from Conus omaria is the only {alpha}-Conotoxin that shows a {approx}20-fold higher affinity to the {alpha}3{beta}2 over the {alpha}6{beta}2 subtype of nicotinic acetylcholine receptor. We have determined a three-dimensional structure of {alpha}-Conotoxin OmIA by nuclear magnetic resonance spectroscopy. {alpha}-Conotoxin OmIA has an '{omega}-shaped' overall topology with His{sup 5}-Asn{sup 12} forming an {alpha}-helix. Structural features of {alpha}-Conotoxin OmIA responsible for its selectivity are suggested by comparing its surface characteristics with other functionally related {alpha}4/7 subfamily Conotoxins. Reduced size of the hydrophilic area in {alpha}-Conotoxin OmIA seems to be associated with the reduced affinity towards the {alpha}6{beta}2 nAChR subtype.
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a novel α Conotoxin peia cloned from conus pergrandis discriminates between rat α9α10 and α7 nicotinic cholinergic receptors
Journal of Biological Chemistry, 2005Co-Authors: Michael J Mcintosh, Baldomero M. Olivera, Maren Watkins, Paola V Plazas, Maria Eugenia Gomezcasati, Belen A ElgoyhenAbstract:The alpha9 and alpha10 nicotinic cholinergic subunits assemble to form the receptor believed to mediate synaptic transmission between efferent olivocochlear fibers and hair cells of the cochlea, one of the few examples of postsynaptic function for a non-muscle nicotinic acetylcholine receptor (nAChR). However, it has been suggested that the expression profile of alpha9 and alpha10 overlaps with that of alpha7 in the cochlea and in sites such as dorsal root ganglion neurons, peripheral blood lymphocytes, developing thymocytes, and skin. We now report the cloning, total synthesis, and characterization of a novel toxin alpha-Conotoxin PeIA that discriminates between alpha9alpha10 and alpha7 nAChRs. This is the first toxin to be identified from Conus pergrandis, a species found in deep waters of the Western Pacific. Alpha-Conotoxin PeIA displayed a 260-fold higher selectivity for alpha-bungarotoxin-sensitive alpha9alpha10 nAChRs compared with alpha-bungarotoxin-sensitive alpha7 receptors. The IC50 of the toxin was 6.9 +/- 0.5 nM and 4.4 +/- 0.5 nM for recombinant alpha9alpha10 and wild-type hair cell nAChRs, respectively. Alpha-Conotoxin PeIA bears high resemblance to alpha-Conotoxins MII and GIC isolated from Conus magus and Conus geographus, respectively. However, neither alpha-Conotoxin MII nor alpha-Conotoxin GIC at concentrations of 10 microM blocked acetylcholine responses elicited in Xenopus oocytes injected with the alpha9 and alpha10 subunits. Among neuronal non-alpha-bungarotoxin-sensitive receptors, alpha-Conotoxin PeIA was also active at alpha3beta2 receptors and chimeric alpha6/alpha3beta2beta3 receptors. Alpha-Conotoxin PeIA represents a novel probe to differentiate responses mediated either through alpha9alpha10 or alpha7 nAChRs in those tissues where both receptors are expressed.
Richard J. Lewis - One of the best experts on this subject based on the ideXlab platform.
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Venom duct origins of prey capture and defensive Conotoxins in piscivorous Conus striatus
'Springer Science and Business Media LLC', 2021Co-Authors: S.w.a. Himaya, Paul Francis Alewood, Ai-hua Jin, Brett Hamilton, Subash K. Rai, Richard J. LewisAbstract:Abstract The venom duct origins of predatory and defensive venoms has not been studied for hook-and-line fish hunting cone snails despite the pharmacological importance of their venoms. To better understand the biochemistry and evolution of injected predatory and defensive venoms, we compared distal, central and proximal venom duct sections across three specimens of C. striatus (Pionoconus) using proteomic and transcriptomic approaches. A total of 370 Conotoxin precursors were identified from the whole venom duct transcriptome. Milked defensive venom was enriched with a potent cocktail of proximally expressed inhibitory α-, ω- and μ-Conotoxins compared to milked predatory venom. In contrast, excitatory κA-Conotoxins dominated both the predatory and defensive venoms despite their distal expression, suggesting this class of Conotoxin can be selectively expressed from the same duct segment in response to either a predatory or defensive stimuli. Given the high abundance of κA-Conotoxins in the Pionoconus clade, we hypothesise that the κA-Conotoxins have evolved through adaptive evolution following their repurposing from ancestral inhibitory A superfamily Conotoxins to facilitate the dietary shift to fish hunting and species radiation in this clade
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Conotoxins: chemistry and biology
Chemical reviews, 2019Co-Authors: Ai-hua Jin, Markus Muttenthaler, Sebastien Dutertre, David J. Craik, Richard J. Lewis, Quentin Kaas, S.w.a. Himaya, Paul Francis AlewoodAbstract:The venom of the marine predatory cone snails (genus Conus) has evolved for prey capture and defense, providing the basis for survival and rapid diversification of the now estimated 750+ species. A typical Conus venom contains hundreds to thousands of bioactive peptides known as Conotoxins. These mostly disulfide-rich and well-structured peptides act on a wide range of targets such as ion channels, G protein-coupled receptors, transporters, and enzymes. Conotoxins are of interest to neuroscientists as well as drug developers due to their exquisite potency and selectivity, not just against prey but also mammalian targets, thereby providing a rich source of molecular probes and therapeutic leads. The rise of integrated venomics has accelerated Conotoxin discovery with now well over 10,000 Conotoxin sequences published. However, their structural and pharmacological characterization lags considerably behind. In this review, we highlight the diversity of new Conotoxins uncovered since 2014, their three-dimensional structures and folds, novel chemical approaches to their syntheses, and their value as pharmacological tools to unravel complex biology. Additionally, we discuss challenges and future directions for the field.
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‘Messy’ Processing of χ-Conotoxin MrIA Generates Homologues with Reduced hNET Potency
MDPI AG, 2019Co-Authors: Rebekah Ziegman, Richard J. Lewis, Andreas Brust, Prerna Jha, Fernanda C. Cardoso, Paul Francis AlewoodAbstract:Integrated venomics techniques have shown that variable processing of Conotoxins from Conus marmoreus resulted in a dramatic expansion in the number of expressed Conotoxins. One Conotoxin from C. marmoreus, the χ-Conotoxin MrIA, is a selective inhibitor of human norepinephrine transporters (hNET) and therefore a drug candidate for attenuating chronic neuropathic pain. It has been found that “messy” processing of the MrIA transcripts results in the expression of MrIA analogs with different truncations of the pro-peptide that contains portions of the MrIA molecule. The aim of this study was to investigate if variable processing of the expressed peptides results in modulation of the existing hNET pharmacology or creates new pharmacologies. To this end, a number of MrIA analogs found in C. marmoreus venom were synthesized and evaluated for their activity at hNET receptors. While several of the analogs exhibited norepinephrine transporter inhibitory activity comparable to that of MrIA, none significantly improved on the potency of Conotoxin MrIA, and those analogs with disrupted pharmacophores produced greatly reduced NET inhibition, confirming previous structure-activity relationships seen on χ-class conopeptides. Additionally, analogs were screened for new activities on ion channels using calcium influx assays, although no major new pharmacology was revealed
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Conotoxin φ mixxviia from the superfamily g2 employs a novel cysteine framework that mimics granulin and displays anti apoptotic activity
Angewandte Chemie, 2017Co-Authors: Zoltan Dekan, Michael J. Smout, Sebastien Dutertre, Norelle L Daly, Alex Loukas, David Wilson, Richard J. Lewis, Paul Francis AlewoodAbstract:Conotoxins are a large family of disulfide-rich peptides that contain unique cysteine frameworks that target a broad range of ion channels and receptors. We recently discovered the 33-residue Conotoxin Φ-MiXXVIIA from Conus miles with a novel cysteine framework comprising three consecutive cysteine residues and four disulfide bonds. Regioselective chemical synthesis helped decipher the disulfide bond connectivity and the structure of Φ-MiXXVIIA was determined by NMR spectroscopy. The 3D structure displays a unique topology containing two β-hairpins that resemble the N-terminal domain of granulin. Similar to granulin, Φ-MiXXVIIA promotes cell proliferation (EC50 17.85 μm) while inhibiting apoptosis (EC50 2.2 μm). Additional framework XXVII sequences were discovered with homologous signal peptides that define the new Conotoxin superfamily G2. The novel structure and biological activity of Φ-MiXXVIIA expands the repertoire of disulfide-rich Conotoxins that recognize mammalian receptors.
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isolation characterization and total regioselective synthesis of the novel μo Conotoxin mfvia from conus magnificus that targets voltage gated sodium channels
Biochemical Pharmacology, 2012Co-Authors: Irina Vetter, Paul Francis Alewood, Zoltan Dekan, David J. Adams, Oliver Knapp, Richard J. LewisAbstract:The μO-Conotoxins are notable for their unique selectivity for Na(v)1.8 over other sodium channel isoforms, making them attractive drug leads for the treatment of neuropathic pain. We describe the discovery of a novel μO-Conotoxin, MfVIA, from the venom of Conus magnificus using high-throughput screening approaches. MfVIA was found to be a hydrophobic 32-residue peptide (amino acid sequence RDCQEKWEYCIVPILGFVYCCPGLICGPFVCV) with highest sequence homology to μO-Conotoxin MrVIB. To overcome the synthetic challenges posed by μO-Conotoxins due to their hydrophobic nature and difficult folding, we developed a novel regioselective approach for the synthesis of μO-Conotoxins. Performing selective oxidative deprotections of the cysteine side-chain protecting groups of the fully protected peptide allowed manipulations in organic solvents with no chromatography required between steps. Using this approach, we obtained correctly folded MfVIA with increased synthetic yields. Biological activity of MfVIA was assessed using membrane potential-sensitive dyes and electrophysiological recording techniques. MfVIA preferentially inhibits Na(v)1.8 (IC₅₀ 95.9±74.3 nM) and Na(v)1.4 (IC₅₀ 81±16 nM), with significantly lower affinity for other Na(v) subtypes (IC₅₀ 431-6203 nM; Na(v)1.5>1.6∼1.7∼1.3∼1.1∼1.2). This improved approach to μO-Conotoxin synthesis will facilitate the optimization of μO-Conotoxins as novel analgesic molecules to improve pain management.
Paul Francis Alewood - One of the best experts on this subject based on the ideXlab platform.
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Venom duct origins of prey capture and defensive Conotoxins in piscivorous Conus striatus
'Springer Science and Business Media LLC', 2021Co-Authors: S.w.a. Himaya, Paul Francis Alewood, Ai-hua Jin, Brett Hamilton, Subash K. Rai, Richard J. LewisAbstract:Abstract The venom duct origins of predatory and defensive venoms has not been studied for hook-and-line fish hunting cone snails despite the pharmacological importance of their venoms. To better understand the biochemistry and evolution of injected predatory and defensive venoms, we compared distal, central and proximal venom duct sections across three specimens of C. striatus (Pionoconus) using proteomic and transcriptomic approaches. A total of 370 Conotoxin precursors were identified from the whole venom duct transcriptome. Milked defensive venom was enriched with a potent cocktail of proximally expressed inhibitory α-, ω- and μ-Conotoxins compared to milked predatory venom. In contrast, excitatory κA-Conotoxins dominated both the predatory and defensive venoms despite their distal expression, suggesting this class of Conotoxin can be selectively expressed from the same duct segment in response to either a predatory or defensive stimuli. Given the high abundance of κA-Conotoxins in the Pionoconus clade, we hypothesise that the κA-Conotoxins have evolved through adaptive evolution following their repurposing from ancestral inhibitory A superfamily Conotoxins to facilitate the dietary shift to fish hunting and species radiation in this clade
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Conotoxins: chemistry and biology
Chemical reviews, 2019Co-Authors: Ai-hua Jin, Markus Muttenthaler, Sebastien Dutertre, David J. Craik, Richard J. Lewis, Quentin Kaas, S.w.a. Himaya, Paul Francis AlewoodAbstract:The venom of the marine predatory cone snails (genus Conus) has evolved for prey capture and defense, providing the basis for survival and rapid diversification of the now estimated 750+ species. A typical Conus venom contains hundreds to thousands of bioactive peptides known as Conotoxins. These mostly disulfide-rich and well-structured peptides act on a wide range of targets such as ion channels, G protein-coupled receptors, transporters, and enzymes. Conotoxins are of interest to neuroscientists as well as drug developers due to their exquisite potency and selectivity, not just against prey but also mammalian targets, thereby providing a rich source of molecular probes and therapeutic leads. The rise of integrated venomics has accelerated Conotoxin discovery with now well over 10,000 Conotoxin sequences published. However, their structural and pharmacological characterization lags considerably behind. In this review, we highlight the diversity of new Conotoxins uncovered since 2014, their three-dimensional structures and folds, novel chemical approaches to their syntheses, and their value as pharmacological tools to unravel complex biology. Additionally, we discuss challenges and future directions for the field.
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‘Messy’ Processing of χ-Conotoxin MrIA Generates Homologues with Reduced hNET Potency
MDPI AG, 2019Co-Authors: Rebekah Ziegman, Richard J. Lewis, Andreas Brust, Prerna Jha, Fernanda C. Cardoso, Paul Francis AlewoodAbstract:Integrated venomics techniques have shown that variable processing of Conotoxins from Conus marmoreus resulted in a dramatic expansion in the number of expressed Conotoxins. One Conotoxin from C. marmoreus, the χ-Conotoxin MrIA, is a selective inhibitor of human norepinephrine transporters (hNET) and therefore a drug candidate for attenuating chronic neuropathic pain. It has been found that “messy” processing of the MrIA transcripts results in the expression of MrIA analogs with different truncations of the pro-peptide that contains portions of the MrIA molecule. The aim of this study was to investigate if variable processing of the expressed peptides results in modulation of the existing hNET pharmacology or creates new pharmacologies. To this end, a number of MrIA analogs found in C. marmoreus venom were synthesized and evaluated for their activity at hNET receptors. While several of the analogs exhibited norepinephrine transporter inhibitory activity comparable to that of MrIA, none significantly improved on the potency of Conotoxin MrIA, and those analogs with disrupted pharmacophores produced greatly reduced NET inhibition, confirming previous structure-activity relationships seen on χ-class conopeptides. Additionally, analogs were screened for new activities on ion channels using calcium influx assays, although no major new pharmacology was revealed
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Conotoxin φ mixxviia from the superfamily g2 employs a novel cysteine framework that mimics granulin and displays anti apoptotic activity
Angewandte Chemie, 2017Co-Authors: Zoltan Dekan, Michael J. Smout, Sebastien Dutertre, Norelle L Daly, Alex Loukas, David Wilson, Richard J. Lewis, Paul Francis AlewoodAbstract:Conotoxins are a large family of disulfide-rich peptides that contain unique cysteine frameworks that target a broad range of ion channels and receptors. We recently discovered the 33-residue Conotoxin Φ-MiXXVIIA from Conus miles with a novel cysteine framework comprising three consecutive cysteine residues and four disulfide bonds. Regioselective chemical synthesis helped decipher the disulfide bond connectivity and the structure of Φ-MiXXVIIA was determined by NMR spectroscopy. The 3D structure displays a unique topology containing two β-hairpins that resemble the N-terminal domain of granulin. Similar to granulin, Φ-MiXXVIIA promotes cell proliferation (EC50 17.85 μm) while inhibiting apoptosis (EC50 2.2 μm). Additional framework XXVII sequences were discovered with homologous signal peptides that define the new Conotoxin superfamily G2. The novel structure and biological activity of Φ-MiXXVIIA expands the repertoire of disulfide-rich Conotoxins that recognize mammalian receptors.
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Comparative Venomics Reveals the Complex Prey Capture Strategy of the Piscivorous Cone Snail Conus catus
Journal of Proteome Research, 2015Co-Authors: S. Himaya, Sebastien Dutertre, Paul Francis Alewood, Ai-hua Jin, Jean Giacomotto, Hoshyar Mohialdeen, Richard LewisAbstract:Venomous marine cone snails produce a unique and remarkably diverse range of venom peptides (Conotoxins and conopeptides) that have proven to be invaluable as pharmacological probes and leads to new therapies. Conus catus is a hook-and-line fish hunter from clade I, with ∼20 Conotoxins identified, including the analgesic ω-Conotoxin CVID (AM336). The current study unravels the venom composition of C. catus with tandem mass spectrometry and 454 sequencing data. From the venom gland transcriptome, 104 precursors were recovered from 11 superfamilies, with superfamily A (especially κA-) Conotoxins dominating (77%) their venom. Proteomic analysis confirmed that κA-Conotoxins dominated the predation-evoked milked venom of each of six C. catus analyzed and revealed remarkable intraspecific variation in both the intensity and type of Conotoxins. High-throughput FLIPR assays revealed that the predation-evoked venom contained a range of Conotoxins targeting the nAChR, Cav, and Nav ion channels, consistent with α- and ω-Conotoxins being used for predation by C. catus. However, the κA-Conotoxins did not act at these targets but induced potent and rapid immobilization followed by bursts of activity and finally paralysis when injected intramuscularly in zebrafish. Our venomics approach revealed the complexity of the envenomation strategy used by C. catus, which contains a mix of both excitatory and inhibitory venom peptides.