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Baldomero M. Olivera - One of the best experts on this subject based on the ideXlab platform.
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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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Conotoxins: from the biodiversity of gastropods to new drugs
Biochemistry (Moscow) Supplement Series B: Biomedical Chemistry, 2012Co-Authors: Alexander E. Fedosov, S. A. Moshkovskii, K. G. Kuznetsova, Baldomero M. OliveraAbstract:The review describes general trends in research of Conotoxins, the peptide toxins isolated from sea gastropods of the genus Conus. It covers publications from conotoxin discovery in 1970th up to contemporary research and considers classification of Conotoxins, their structural diversity and different ways of their action on molecular targets, particularly, ion channels. Special attention is paid to the applied aspect of conotoxin research especially to perspectives of the development of conotoxin based drugs. The first example of such conotoxin based drug is ziconotide an analgesic of a new generation.
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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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conus venoms a rich source of peptide based therapeutics
Current Pharmaceutical Design, 2008Co-Authors: Tiffany S Han, Baldomero M. Olivera, Russell W. Teichert, Grzegorz BulajAbstract:Over two decades of research on venom peptides derived from cone snails (“conopeptides or Conotoxins”) has led to several compounds that have reached human clinical trials, most of them for the treatment of pain. Remarkably, none of the conopeptides in clinical development mediate analgesia through the opioid receptors, underlying the diverse and novel neuropharmacology evolved by Conus snails. These predatory animals produce an estimated ∼100,000 distinct Conotoxins, a vast majority yet to be discovered and characterized. The conopeptides studied to-date in animal models, have exhibited antinociceptive, antiepileptic, neuroprotective or cardioprotective activities. Screening results also suggest applications of Conotoxins in cancer, neuromuscular and psychiatric disorders. Additional potentially important applications of conotoxin research are the discovery and validation of new therapeutic targets, also defining novel binding sites on already validated molecular targets. As the structural and functional diversity of Conotoxins is being investigated, the Conus venoms continue to surprise with the plethora of neuropharmacological compounds and potential new therapeutics. This review summarizes recent efforts in the discovery of conopeptides, and their preclinical and clinical development.
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muo Conotoxins inhibit nav channels by interfering with their voltage sensors in domain 2
Channels, 2007Co-Authors: Enrico Leipold, Baldomero M. Olivera, Heinrich Terlau, Herbert Debie, Stefan Zorn, Borges Adolfo, Stefan H HeinemannAbstract:The µO-Conotoxins MrVIA and MrVIB are 31-residue peptides from Conus marmoreus, belonging to the O-superfamily of Conotoxins with three disulfide bridges. They have attracted attention because they are inhibitors of tetrodotoxin-insensitive voltage-gated sodium channels (NaV1.8) and could therefore serve as lead structure for novel analgesics. The aim of this study was to elucidate the molecular mechanism by which µO-Conotoxins affect NaV channels. Rat NaV1.4 channels and mutants thereof were expressed in mammalian cells and were assayed with the whole-cell patch-clamp method. Unlike for the M-superfamily µ-conotoxin GIIIA from Conus geographus, channel block by MrVIA was strongly diminished after activating the NaV channels by depolarizing voltage steps. Searching for the source of this voltage dependence, the gating charges in all four voltage sensors were reduced by site-directed mutagenesis showing that alterations of the voltage sensor in domain-2 have the strongest impact on MrVIA action. These resu...
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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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.
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molecular engineering of Conotoxins the importance of loop size to α conotoxin structure and function
Journal of Medicinal Chemistry, 2008Co-Authors: Norelle L Daly, Sebastien Dutertre, Simon T. Nevin, David J. Craik, Richard J. Lewis, David J. Adams, Chingi Anderson Wang, Paul F. AlewoodAbstract:α-Conotoxins are competitive antagonists of nicotinic acetylcholine receptors (nAChRs). The majority of currently characterized α-Conotoxins have a 4/7 loop size, and the major features of neuronal α-Conotoxins include a globular disulfide connectivity and a helical structure centered around the third of their four cysteine residues. In this study, a novel “molecular pruning” approach was undertaken to define the relationship between loop size, structure, and function of α-Conotoxins. This involved the systematic truncation of the second loop in the α-conotoxin [A10L]PnIA [4/7], a potent antagonist of the α7 nAChR. The penalty for truncation was found to be decreased conformational stability and increased susceptibility to disulfide bond scrambling. Truncation down to 4/4[A10L]PnIA maintained helicity and did not significantly reduce electrophysiological activity at α7 nAChRs, whereas 4/3[A10L]PnIA lost both α7 nAChR activity and helicity. In contrast, all truncated analogues lost ∼100-fold affinity at th...
Qiong Shi - One of the best experts on this subject based on the ideXlab platform.
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High-Throughput Identification and Analysis of Novel Conotoxins from Three Vermivorous Cone Snails by Transcriptome Sequencing
MDPI AG, 2019Co-Authors: Ge Yao, Hui Jiang, Chao Peng, Yabing Zhu, Chongxu Fan, Jisheng Chen, Ying Cao, Qiong ShiAbstract:The venom of each Conus species consists of a diverse array of neurophysiologically active peptides, which are mostly unique to the examined species. In this study, we performed high-throughput transcriptome sequencing to extract and analyze putative conotoxin transcripts from the venom ducts of 3 vermivorous cone snails (C. caracteristicus, C. generalis, and C. quercinus), which are resident in offshore waters of the South China Sea. In total, 118, 61, and 48 putative Conotoxins (across 22 superfamilies) were identified from the 3 Conus species, respectively; most of them are novel, and some possess new cysteine patterns. Interestingly, a series of 45 unassigned Conotoxins presented with a new framework of C-C-C-C-C-C, and their mature regions were sufficiently distinct from any other known Conotoxins, most likely representing a new superfamily. O- and M-superfamily Conotoxins were the most abundant in transcript number and transcription level, suggesting their critical roles in the venom functions of these vermivorous cone snails. In addition, we identified numerous functional proteins with potential involvement in the biosynthesis, modification, and delivery process of Conotoxins, which may shed light on the fundamental mechanisms for the generation of these important Conotoxins within the venom duct of cone snails
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high throughput identification of novel Conotoxins from the vermivorous oak cone snail conus quercinus by transcriptome sequencing
International Journal of Molecular Sciences, 2018Co-Authors: Bingmiao Gao, Chao Peng, Yabing Zhu, Yuhui Sun, Tian Zhao, Yu Huang, Qiong ShiAbstract:The primary objective of this study was to realize the large-scale discovery of conotoxin sequences from different organs (including the venom duct, venom bulb and salivary gland) of the vermivorous Oak cone snail, Conus quercinus. Using high-throughput transcriptome sequencing, we identified 133 putative Conotoxins that belong to 34 known superfamilies, of which nine were previously reported while the remaining 124 were novel Conotoxins, with 17 in new and unassigned conotoxin groups. A-, O1-, M-, and I2- superfamilies were the most abundant, and the cysteine frameworks XIII and VIII were observed for the first time in the A- and I2-superfamilies. The transcriptome data from the venom duct, venom bulb and salivary gland showed considerable inter-organizational variations. Each organ had many exclusive Conotoxins, and only seven of all the inferred mature peptides were common in the three organs. As expected, most of the identified Conotoxins were synthesized in the venom duct at relatively high levels; however, a number of Conotoxins were also identified in the venom bulb and the salivary gland with very low transcription levels. Therefore, various organs have different Conotoxins with high diversity, suggesting greater contributions from several organs to the high-throughput discovery of new Conotoxins for future drug development.
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cone snails a big store of Conotoxins for novel drug discovery
Toxins, 2017Co-Authors: Bingmiao Gao, Chao Peng, Jiaan Yang, Junqing Zhang, Qiong ShiAbstract:Marine drugs have developed rapidly in recent decades. Cone snails, a group of more than 700 species, have always been one of the focuses for new drug discovery. These venomous snails capture prey using a diverse array of unique bioactive neurotoxins, usually named as Conotoxins or conopeptides. These Conotoxins have proven to be valuable pharmacological probes and potential drugs due to their high specificity and affinity to ion channels, receptors, and transporters in the nervous systems of target prey and humans. Several research groups, including ours, have examined the venom gland of cone snails using a combination of transcriptomic and proteomic sequencing, and revealed the existence of hundreds of conotoxin transcripts and thousands of conopeptides in each Conus species. Over 2000 nucleotide and 8000 peptide sequences of Conotoxins have been published, and the number is still increasing quickly. However, more than 98% of these sequences still lack 3D structural and functional information. With the rapid development of genomics and bioinformatics in recent years, functional predictions and investigations on Conotoxins are making great progress in promoting the discovery of novel drugs. For example, ω-MVIIA was approved by the U.S. Food and Drug Administration in 2004 to treat chronic pain, and nine more Conotoxins are at various stages of preclinical or clinical evaluation. In short, the genus Conus, the big family of cone snails, has become an important genetic resource for conotoxin identification and drug development.
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a transcriptomic survey of ion channel based Conotoxins in the chinese tubular cone snail conus betulinus
Marine Drugs, 2017Co-Authors: Yu Huang, Bingmiao Gao, Chao Peng, Qiong ShiAbstract:Conotoxins in the venom of cone snails (Conus spp.) are a mixture of active peptides that work as blockers, agonists, antagonists, or inactivators of various ion channels. Recently we reported a high-throughput method to identify 215 conotoxin transcripts from the Chinese tubular cone snail, C. betulinus. Here, based on the previous datasets of four transcriptomes from three venom ducts and one venom bulb, we explored ion channel-based Conotoxins and predicted their related ion channel receptors. Homologous analysis was also performed for the most abundant ion channel protein, voltage-gated potassium (Kv; with Kv1.1 as the representative), and the most studied ion channel receptor, nicotinic acetylcholine receptor (nAChR; with α2-nAChR as the representative), in different animals. Our transcriptomic survey demonstrated that ion channel-based Conotoxins and related ion channel proteins/receptors transcribe differentially between the venom duct and the venom bulb. In addition, we observed that putative κ-Conotoxins were the most common Conotoxins with the highest transcription levels in the examined C. betulinus. Furthermore, Kv1.1 and α2-nAChR were conserved in their functional domains of deduced protein sequences, suggesting similar effects of Conotoxins via the ion channels in various species, including human beings. In a word, our present work suggests a high-throughput way to develop Conotoxins as potential drugs for treatment of ion channel-associated human diseases.
Richard J. Lewis - One of the best experts on this subject based on the ideXlab platform.
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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 F. 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 F. 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, Irina Vetter, 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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The role of defensive ecological interactions in the evolution of Conotoxins
Molecular ecology, 2016Co-Authors: Jutty Rajan Prashanth, Sebastien Dutertre, Ai-hua Jin, Paul F. Alewood, Fernanda C. Cardoso, V. Lavergne, B. Hamilton, J. Griffin, Deon J. Venter, Richard J. LewisAbstract:Venoms comprise of complex mixtures of peptides evolved for predation and defensive purposes. Remarkably, some carnivorous cone snails can inject two distinct venoms in response to predatory or defensive stimuli, providing a unique opportunity to study separately how different ecological pressures contribute to toxin diversification. Here, we report the extraordinary defensive strategy of the Rhizoconus subgenus of cone snails. The defensive venom from this worm-hunting subgenus is unusually simple, almost exclusively composed of αD-Conotoxins instead of the ubiquitous αA-Conotoxins found in the more complex defensive venom of mollusc- and fish-hunting cone snails. A similarly compartmentalized venom gland as those observed in the other dietary groups facilitates the deployment of this defensive venom. Transcriptomic analysis of a Conus vexillum venom gland revealed the αD-Conotoxins as the major transcripts, with lower amounts of 15 known and four new conotoxin superfamilies also detected with likely roles in prey capture. Our phylogenetic and molecular evolution analysis of the αD-Conotoxins from five subgenera of cone snails suggests they evolved episodically as part of a defensive strategy in the Rhizoconus subgenus. Thus, our results demonstrate an important role for defence in the evolution of Conotoxins.
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solving the α conotoxin folding problem efficient selenium directed on resin generation of more potent and stable nicotinic acetylcholine receptor antagonists
Journal of the American Chemical Society, 2010Co-Authors: Markus Muttenthaler, Alexander A. Grishin, P T Choy, Christopher J Armishaw, Ci Anderson Wang, Norelle L Daly, Shuhong Hu, Simon T. Nevin, Richard J. Lewis, Jennifer L. MartinAbstract:α-Conotoxins are tightly folded miniproteins that antagonize nicotinic acetylcholine receptors (nAChR) with high specificity for diverse subtypes. Here we report the use of selenocysteine in a supported phase method to direct native folding and produce α-Conotoxins efficiently with improved biophysical properties. By replacing complementary cysteine pairs with selenocysteine pairs on an amphiphilic resin, we were able to chemically direct all five structural subclasses of α-Conotoxins exclusively into their native folds. X-ray analysis at 1.4 A resolution of α-selenoconotoxin PnIA confirmed the isosteric character of the diselenide bond and the integrity of the α-conotoxin fold. The α-selenoConotoxins exhibited similar or improved potency at rat diaphragm muscle and α3β4, α7, and α1β1δγ nAChRs expressed in Xenopus oocytes plus improved disulfide bond scrambling stability in plasma. Together, these results underpin the development of more stable and potent nicotinic antagonists suitable for new drug therap...
Grzegorz Bulaj - One of the best experts on this subject based on the ideXlab platform.
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structure and function of μ Conotoxins peptide based sodium channel blockers with analgesic activity
Future Medicinal Chemistry, 2014Co-Authors: Brad Reed Green, Grzegorz Bulaj, Raymond S. NortonAbstract:μ-Conotoxins block voltage-gated sodium channels (VGSCs) and compete with tetrodotoxin for binding to the sodium conductance pore. Early efforts identified µ-Conotoxins that preferentially blocked the skeletal muscle subtype (NaV1.4). However, the last decade witnessed a significant increase in the number of µ-Conotoxins and the range of VGSC subtypes inhibited (NaV1.2, NaV1.3 or NaV1.7). Twenty µ-conotoxin sequences have been identified to date and structure–activity relationship studies of several of these identified key residues responsible for interactions with VGSC subtypes. Efforts to engineer-in subtype specificity are driven by in vivo analgesic and neuromuscular blocking activities. This review summarizes structural and pharmacological studies of µ-Conotoxins, which show promise for development of selective blockers of NaV1.2, and perhaps also NaV1.1,1.3 or 1.7.
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conus venoms a rich source of peptide based therapeutics
Current Pharmaceutical Design, 2008Co-Authors: Tiffany S Han, Baldomero M. Olivera, Russell W. Teichert, Grzegorz BulajAbstract:Over two decades of research on venom peptides derived from cone snails (“conopeptides or Conotoxins”) has led to several compounds that have reached human clinical trials, most of them for the treatment of pain. Remarkably, none of the conopeptides in clinical development mediate analgesia through the opioid receptors, underlying the diverse and novel neuropharmacology evolved by Conus snails. These predatory animals produce an estimated ∼100,000 distinct Conotoxins, a vast majority yet to be discovered and characterized. The conopeptides studied to-date in animal models, have exhibited antinociceptive, antiepileptic, neuroprotective or cardioprotective activities. Screening results also suggest applications of Conotoxins in cancer, neuromuscular and psychiatric disorders. Additional potentially important applications of conotoxin research are the discovery and validation of new therapeutic targets, also defining novel binding sites on already validated molecular targets. As the structural and functional diversity of Conotoxins is being investigated, the Conus venoms continue to surprise with the plethora of neuropharmacological compounds and potential new therapeutics. This review summarizes recent efforts in the discovery of conopeptides, and their preclinical and clinical development.
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biochemical and gene expression analyses of Conotoxins in conus textile venom ducts
Biochemical and Biophysical Research Communications, 2005Co-Authors: James E Garrett, Baldomero M. Olivera, Maren Watkins, Olga Buczek, Grzegorz BulajAbstract:Each Conus snail species produces 50-200 unique peptide-based Conotoxins, derived from a number of different gene superfamilies. Conotoxins are synthesized and secreted in a long venom duct, but biochemical and molecular aspects of their biosynthesis remain poorly understood. Here, we analyzed expression patterns of conotoxin genes belonging to different superfamilies in Conus textile venom ducts. The results demonstrate that specific gene families are expressed in particular regions of the venom duct. Biochemical analysis using liquid chromatography and mass spectrometry revealed an even more localized accumulation of individual Conotoxins. This study demonstrates for the first time that specialization of gene expression, processing, and secretion of Conotoxins occurs in different regions of the venom duct.
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αA-Conotoxin OIVA defines a new αA-conotoxin subfamily of nicotinic acetylcholine receptor inhibitors
Toxicon : official journal of the International Society on Toxinology, 2004Co-Authors: Russell W. Teichert, Jean Rivier, John Dykert, Laura A. Cervini, Jozsef Gulyas, Grzegorz Bulaj, Michael Ellison, Baldomero M. OliveraAbstract:Abstract The venoms of cone snails are rich in multiply disulfide-crosslinked peptides, the Conotoxins. Conotoxins are grouped into families on the basis of shared cysteine patterns and homologous molecular targets. For example, both the κA- and αA-conotoxin families share the same Class IV Cys pattern (–CC–C–C–C–C–), but differ in their molecular targets. The κA-Conotoxins are excitatory toxins that purportedly block potassium channels, while the αA-Conotoxins are paralytic Conotoxins that inhibit nicotinic acetylcholine receptors (nAChRs). In this work, we describe the isolation and characterization of a novel Conus peptide from venom milked from Hawaiian specimens of Conus obscurus . This peptide shares the Class IV Cys pattern but differs from both previously characterized αA- and κA-Conotoxins in the spacing of amino acids between Cys resides. However, the peptide is similar to previously characterized αA-Conotoxins in its paralytic effects on fish and its antagonist activity on the neuromuscular nAChR. Unexpectedly, the peptide differs in its disulfide bonding from αA-conotoxin PIVA. We have named this unique peptide αA-conotoxin OIVA, and we consider it the defining member of a subfamily of αA-Conotoxins that we designate the αA (1–3) -Conotoxins to identify them by their unique disulfide bonding framework. These results indicate that the αA-conotoxin family is both more structurally diverse and broadly distributed than previously believed.
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δ-Conotoxin Structure/Function through a Cladistic Analysis†
Biochemistry, 2001Co-Authors: Grzegorz Bulaj, Doju Yoshikami, Maren Watkins, Richard G. Delacruz, Aryan Azimi-zonooz, Peter J. West, Baldomero M. OliveraAbstract:δ-Conotoxins are Conus peptides that inhibit inactivation of voltage-gated sodium channels. The suggestion that δ-Conotoxins might be an essential component of the venoms of fish-hunting cone snails which rapidly immobilize their prey [Terlau, H., Shon, K., Grilley, M., Stocker, M., Stuhmer, W., and Olivera, B. M. (1996) Nature 381, 148−151] has not been tested. On the basis of cDNA cloning, all of the fish-hunting Conus analyzed yielded at least one δ-conotoxin sequence. In addition, one δ-conotoxin isolated from the venom of Conus striatus had an amino acid sequence identical to that predicted from cDNA cloning. This new peptide exhibited properties of δ-Conotoxins: it targeted sodium channels and potentiated action potentials by slowing channel inactivation. Homologous sequences of δ-Conotoxins from two groups (clades) of related fish-hunting Conus species share consensus features but differ significantly from the two known δ-Conotoxins from mollusc-hunting Conus venoms. Three large hydrophobic amino ...