scholarly journals Solution conformation of alpha-conotoxin GIC, a novel potent antagonist of alpha3beta2 nicotinic acetylcholine receptors

2004 ◽  
Vol 380 (2) ◽  
pp. 347-352 ◽  
Author(s):  
Seung-Wook CHI ◽  
Do-Hyoung KIM ◽  
Baldomero M. OLIVERA ◽  
J. Michael McINTOSH ◽  
Kyou-Hoon HAN
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Solution Structure ◽  
Heavy Atom ◽  
Three Dimensional ◽  
Cone Snail ◽  
Dimensional Solution ◽  
Nicotinic Acetylcholine ◽  
Binding Characteristics ◽  
Molecular Features

α-Conotoxin GIC is a 16-residue peptide isolated from the venom of the cone snail Conus geographus. α-Conotoxin GIC potently blocks the α3β2 subtype of human nicotinic acetylcholine receptor, showing a high selectivity for neuronal versus muscle subtype [McIntosh, Dowell, Watkins, Garrett, Yoshikami, and Olivera (2002) J. Biol. Chem. 277, 33610–33615]. We have now determined the three-dimensional solution structure of α-conotoxin GIC by NMR spectroscopy. The structure of α-conotoxin GIC is well defined with backbone and heavy atom root mean square deviations (residues 2–16) of 0.53 Å and 0.96 Å respectively. Structure and surface comparison of α-conotoxin GIC with the other α4/7 subfamily conotoxins reveals unique structural aspects of α-conotoxin GIC. In particular, the structural comparison between α-conotoxins GIC and MII indicates molecular features that may confer their similar receptor specificity profile, as well as those that provide the unique binding characteristics of α-conotoxin GIC.

scholarly journals Ribbon α-Conotoxin KTM Exhibits Potent Inhibition of Nicotinic Acetylcholine Receptors

Marine Drugs ◽  
2019 ◽  
Vol 17 (12) ◽  
pp. 669
Author(s):  
Leanna A. Marquart ◽  
Matthew W. Turner ◽  
Lisa R. Warner ◽  
Matthew D. King ◽  
James R. Groome ◽  
...  
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Nicotinic Acetylcholine ◽  
Amino Acid Peptide ◽  
Potent Inhibition ◽  
Helical Content ◽  
Distance Restraints ◽  
Native Fold

KTM is a 16 amino acid peptide with the sequence WCCSYPGCYWSSSKWC. Here, we present the nuclear magnetic resonance (NMR) structure and bioactivity of this rationally designed α-conotoxin (α-CTx) that demonstrates potent inhibition of rat α3β2-nicotinic acetylcholine receptors (rα3β2-nAChRs). Two bioassays were used to test the efficacy of KTM. First, a qualitative PC12 cell-based assay confirmed that KTM acts as a nAChR antagonist. Second, bioactivity evaluation by two-electrode voltage clamp electrophysiology was used to measure the inhibition of rα3β2-nAChRs by KTM (IC50 = 0.19 ± 0.02 nM), and inhibition of the same nAChR isoform by α-CTx MII (IC50 = 0.35 ± 0.8 nM). The three-dimensional structure of KTM was determined by NMR spectroscopy, and the final set of 20 structures derived from 32 distance restraints, four dihedral angle constraints, and two disulfide bond constraints overlapped with a mean global backbone root-mean-square deviation (RMSD) of 1.7 ± 0.5 Å. The structure of KTM did not adopt the disulfide fold of α-CTx MII for which it was designed, but instead adopted a flexible ribbon backbone and disulfide connectivity of C2–C16 and C3–C8 with an estimated 12.5% α-helical content. In contrast, α-CTx MII, which has a native fold of C2–C8 and C3–C16, has an estimated 38.1% α-helical secondary structure. KTM is the first reported instance of a Framework I (CC-C-C) α-CTx with ribbon connectivity to display sub-nanomolar inhibitory potency of rα3β2-nAChR subtypes.

scholarly journals αM-Conotoxin MIIIJ Blocks Nicotinic Acetylcholine Receptors at Neuromuscular Junctions of Frog and Fish

Toxins ◽  
2020 ◽  
Vol 12 (3) ◽  
pp. 197
Author(s):  
Matthew J. Rybin ◽  
Henrik O’Brien ◽  
Iris Bea L. Ramiro ◽  
Layla Azam ◽  
J. Michael McIntosh ◽  
...  
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Neuromuscular Junctions ◽  
Functional Characterization ◽  
Cone Snail ◽  
Nicotinic Acetylcholine ◽  
New Class ◽  
Goldfish Carassius Auratus ◽  
Snake Neurotoxin

We report the discovery and functional characterization of αM-Conotoxin MIIIJ, a peptide from the venom of the fish-hunting cone snail Conus magus. Injections of αM-MIIIJ induced paralysis in goldfish (Carassius auratus) but not mice. Intracellular recording from skeletal muscles of fish (C. auratus) and frog (Xenopus laevis) revealed that αM-MIIIJ inhibited postsynaptic nicotinic acetylcholine receptors (nAChRs) with an IC50 of ~0.1 μM. With comparable potency, αM-MIIIJ reversibly blocked ACh-gated currents (IACh) of voltage-clamped X. laevis oocytes exogenously expressing nAChRs cloned from zebrafish (Danio rerio) muscle. αM-MIIIJ also protected against slowly-reversible block of IACh by α-bungarotoxin (α-BgTX, a snake neurotoxin) and α-conotoxin EI (α-EI, from Conus ermineus another fish hunter) that competitively block nAChRs at the ACh binding site. Furthermore, assessment by fluorescence microscopy showed that αM-MIIIJ inhibited the binding of fluorescently-tagged α-BgTX at neuromuscular junctions of X. laevis, C. auratus, and D. rerio. (Note, we observed that αM-MIIIJ can block adult mouse and human muscle nAChRs exogenously expressed in X. laevis oocytes, but with IC50s ~100-times higher than those of zebrafish nAChRs.) Taken together, these results indicate that αM-MIIIJ inhibits muscle nAChRs and furthermore apparently does so by interfering with the binding of ACh to its receptor. Comparative alignments with homologous sequences identified in other fish hunters revealed that αM-MIIIJ defines a new class of muscle nAChR inhibitors from cone snails.

Solution structure of α-conotoxin PIA, a novel antagonist of α6 subunit containing nicotinic acetylcholine receptors

2005 ◽  
Vol 338 (4) ◽  
pp. 1990-1997 ◽  
Author(s):  
Seung-Wook Chi ◽  
Si-Hyung Lee ◽  
Do-Hyoung Kim ◽  
Jae-Sung Kim ◽  
Baldomero M. Olivera ◽  
...  
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Solution Structure ◽  
Nicotinic Acetylcholine

Functional determinants by which snake and cone snail toxins block the α7 neuronal nicotinic acetylcholine receptors

1998 ◽  
Vol 92 (2) ◽  
pp. 107-111 ◽  
Author(s):  
Denis Servent ◽  
Hung Lam Thanh ◽  
Stéphanie Antil ◽  
Daniel Bertrand ◽  
Pierre-Jean Corringer ◽  
...  
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Neuronal Nicotinic Acetylcholine Receptors ◽  
Cone Snail ◽  
Nicotinic Acetylcholine

Three-Dimensional Solution Structure of α-Conotoxin MII, an α3β2Neuronal Nicotinic Acetylcholine Receptor-Targeted Ligand†,‡

Biochemistry ◽  
1997 ◽  
Vol 36 (50) ◽  
pp. 15693-15700 ◽  
Author(s):  
Ki-Joon Shon ◽  
Steven C. Koerber ◽  
Jean E. Rivier ◽  
Baldomero M. Olivera ◽  
J. Michael McIntosh
Keyword(s):  
Acetylcholine Receptor ◽  
Nicotinic Acetylcholine Receptor ◽  
Solution Structure ◽  
Three Dimensional ◽  
Dimensional Solution ◽  
Nicotinic Acetylcholine

scholarly journals Constructing and Tuning Excitatory Cholinergic Synapses: The Multifaceted Functions of Nicotinic Acetylcholine Receptors in Drosophila Neural Development and Physiology

2021 ◽  
Vol 15 ◽  
Author(s):  
Justin S. Rosenthal ◽  
Quan Yuan
Keyword(s):  
Nervous System ◽  
Nicotinic Acetylcholine Receptors ◽  
Neural Development ◽  
Acetylcholine Receptors ◽  
Animal Studies ◽  
Neuromuscular Junctions ◽  
Nicotinic Acetylcholine ◽  
Molecular Features ◽  
Cholinergic Synapses

Nicotinic acetylcholine receptors (nAchRs) are widely distributed within the nervous system across most animal species. Besides their well-established roles in mammalian neuromuscular junctions, studies using invertebrate models have also proven fruitful in revealing the function of nAchRs in the central nervous system. During the earlier years, both in vitro and animal studies had helped clarify the basic molecular features of the members of the Drosophila nAchR gene family and illustrated their utility as targets for insecticides. Later, increasingly sophisticated techniques have illuminated how nAchRs mediate excitatory neurotransmission in the Drosophila brain and play an integral part in neural development and synaptic plasticity, as well as cognitive processes such as learning and memory. This review is intended to provide an updated survey of Drosophila nAchR subunits, focusing on their molecular diversity and unique contributions to physiology and plasticity of the fly neural circuitry. We will also highlight promising new avenues for nAchR research that will likely contribute to better understanding of central cholinergic neurotransmission in both Drosophila and other organisms.

scholarly journals Concatenated nicotinic acetylcholine receptors: A gift or a curse?

2018 ◽  
Vol 150 (3) ◽  
pp. 453-473 ◽  
Author(s):  
Philip Kiær Ahring ◽  
Vivian Wan Yu Liao ◽  
Thomas Balle
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Three Dimensional ◽  
Mammalian Brain ◽  
Nicotinic Acetylcholine ◽  
Novel Treatments ◽  
Full Control ◽  
Ion Conducting ◽  
Receptor Assembly ◽  
Receptor Family

Nicotinic acetylcholine receptors (nAChRs) belong to the Cys-loop receptor family and are vital for normal mammalian brain function. Cys-loop receptors are pentameric ligand-gated ion channels formed from five identical or homologous subunits oriented around a central ion-conducting pore, which result in homomeric or heteromeric receptors, respectively. Within a given Cys-loop receptor family, many different heteromeric receptors can assemble from a common set of subunits, and understanding the properties of these heteromeric receptors is crucial for the continuing quest to generate novel treatments for human diseases. Yet this complexity also presents a hindrance for studying Cys-loop receptors in heterologous expression systems, where full control of the receptor stoichiometry and assembly is required. Therefore, subunit concatenation technology is commonly used to control receptor assembly. In theory, this methodology should facilitate full control of the stoichiometry. In reality, however, we find that commonly used constructs do not yield the expected receptor stoichiometries. With ternary or more complex receptors, concatenated subunits must assemble uniformly in only one orientation; otherwise, the resulting receptor pool will consist of receptors with mixed stoichiometries. We find that typically used constructs of α4β2 nAChR dimers, tetramers, and pentamers assemble readily in both the clockwise and the counterclockwise orientations. Consequently, we investigate the possibility of successfully directing the receptor assembly process using concatenation. We begin by investigating the three-dimensional structures of the α4β2 nAChR. Based on this, we hypothesize that the minimum linker length required to bridge the C terminus of one subunit to the N terminus of the next is shortest in the counterclockwise orientation. We then successfully express receptors with a uniform stoichiometry by systematically shortening linker lengths, proving the hypothesis correct. Our results will significantly aid future studies of heteromeric Cys-loop receptors and enable clarification of the current contradictions in the literature.

scholarly journals Structural Diversity and Dynamics of Human Three-Finger Proteins Acting on Nicotinic Acetylcholine Receptors

2020 ◽  
Vol 21 (19) ◽  
pp. 7280
Author(s):  
Alexander S. Paramonov ◽  
Milita V. Kocharovskaya ◽  
Andrey V. Tsarev ◽  
Dmitrii S. Kulbatskii ◽  
Eugene V. Loktyushov ◽  
...  
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
3D Structure ◽  
Three Dimensional ◽  
Structural Diversity ◽  
Backbone Dynamics ◽  
Surface Distribution ◽  
Nicotinic Acetylcholine ◽  
Conformational Plasticity ◽  
Β Sheet

Ly-6/uPAR or three-finger proteins (TFPs) contain a disulfide-stabilized β-structural core and three protruding loops (fingers). In mammals, TFPs have been found in epithelium and the nervous, endocrine, reproductive, and immune systems. Here, using heteronuclear NMR, we determined the three-dimensional (3D) structure and backbone dynamics of the epithelial secreted protein SLURP-1 and soluble domains of GPI-anchored TFPs from the brain (Lynx2, Lypd6, Lypd6b) acting on nicotinic acetylcholine receptors (nAChRs). Results were compared with the data about human TFPs Lynx1 and SLURP-2 and snake α-neurotoxins WTX and NTII. Two different topologies of the β-structure were revealed: one large antiparallel β-sheet in Lypd6 and Lypd6b, and two β-sheets in other proteins. α-Helical segments were found in the loops I/III of Lynx2, Lypd6, and Lypd6b. Differences in the surface distribution of charged and hydrophobic groups indicated significant differences in a mode of TFPs/nAChR interactions. TFPs showed significant conformational plasticity: the loops were highly mobile at picosecond-nanosecond timescale, while the β-structural regions demonstrated microsecond-millisecond motions. SLURP-1 had the largest plasticity and characterized by the unordered loops II/III and cis-trans isomerization of the Tyr39-Pro40 bond. In conclusion, plasticity could be an important feature of TFPs adapting their structures for optimal interaction with the different conformational states of nAChRs.

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