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 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 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.

scholarly journals Synthesis, Structural and Pharmacological Characterizations of CIC, a Novel α-Conotoxin with an Extended N-terminal Tail

Marine Drugs ◽  
2021 ◽  
Vol 19 (3) ◽  
pp. 141
Author(s):  
Julien Giribaldi ◽  
Yves Haufe ◽  
Edward R. J. Evans ◽  
David T. Wilson ◽  
Norelle L. Daly ◽  
...  
Keyword(s):  
Biological Activity ◽  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Three Dimensional ◽  
Venom Gland ◽  
Dimensional Structure ◽  
Major Influence ◽  
Dominant Group ◽  
Marine Predators ◽  
Pharmacological Targets

Cone snails are venomous marine predators that rely on fast-acting venom to subdue their prey and defend against aggressors. The conotoxins produced in the venom gland are small disulfide-rich peptides with high affinity and selectivity for their pharmacological targets. A dominant group comprises α-conotoxins, targeting nicotinic acetylcholine receptors. Here, we report on the synthesis, structure determination and biological activity of a novel α-conotoxin, CIC, found in the predatory venom of the piscivorous species Conus catus and its truncated mutant Δ-CIC. CIC is a 4/7 α-conotoxin with an unusual extended N-terminal tail. High-resolution NMR spectroscopy shows a major influence of the N-terminal tail on the apparent rigidity of the three-dimensional structure of CIC compared to the more flexible Δ-CIC. Surprisingly, this effect on the structure does not alter the biological activity, since both peptides selectively inhibit α3β2 and α6/α3β2β3 nAChRs with almost identical sub- to low micromolar inhibition constants. Our results suggest that the N-terminal part of α-conotoxins can accommodate chemical modifications without affecting their pharmacology.

scholarly journals The 3/4- and 3/6-Subfamily Variants of α-Conotoxins GI and MI Exhibit Potent Inhibitory Activity against Muscular Nicotinic Acetylcholine Receptors

Marine Drugs ◽  
2021 ◽  
Vol 19 (12) ◽  
pp. 705
Author(s):  
Xiaoli Ma ◽  
Qiuyuan Huang ◽  
Shuo Yu ◽  
Shujing Xu ◽  
Yue Huang ◽  
...  
Keyword(s):  
Disulfide Bond ◽  
Nicotinic Acetylcholine Receptors ◽  
Disulfide Bonds ◽  
Acetylcholine Receptors ◽  
Basic Amino Acid ◽  
Wild Type ◽  
Nicotinic Acetylcholine ◽  
Potent Inhibition ◽  
Potent Inhibitory Activity

α-Conotoxins GI and MI belong to the 3/5 subfamily of α-conotoxins and potently inhibit muscular nicotinic acetylcholine receptors (nAChRs). To date, no 3/4- or 3/6-subfamily α-conotoxins have been reported to inhibit muscular nAChRs. In the present study, a series of new 3/4-, 3/6-, and 3/7-subfamily GI and MI variants were synthesized and functionally characterized by modifications of loop2. The results show that the 3/4-subfamily GI variant GI[∆8G]-II and the 3/6-subfamily variants GI[+13A], GI[+13R], and GI[+13K] displayed potent inhibition of muscular nAChRs expressed in Xenopus oocytes, with an IC50 of 45.4–73.4 nM, similar to or slightly lower than that of wild-type GI (42.0 nM). The toxicity of these GI variants in mice appeared to be about a half to a quarter of that of wild-type GI. At the same time, the 3/7-subfamily GI variants showed significantly lower in vitro potency and toxicity. On the other hand, similar to the 3/6-subfamily GI variants, the 3/6-subfamily MI variants MI[+14R] and MI[+14K] were also active after the addition of a basic amino acid, Arg or Lys, in loop2, but the activity was not maintained for the 3/4-subfamily MI variant MI [∆9G]. Interestingly, the disulfide bond connectivity “C1–C4, C2–C3” in the 3/4-subfamily variant GI[∆8G]-II was significantly more potent than the “C1–C3, C2–C4” connectivity found in wild-type GI and MI, suggesting that disulfide bond connectivity is easily affected in the rigid 3/4-subfamily α-conotoxins and that the disulfide bonds significantly impact the variants’ function. This work is the first to demonstrate that 3/4- and 3/6-subfamily α-conotoxins potently inhibit muscular nAChRs, expanding our knowledge of α-conotoxins and providing new motifs for their further modifications.

Nicotinic acetylcholine receptors in the patients with Parkinson's disease: 5IA-SPECT

2005 ◽  
Vol 25 (1_suppl) ◽  
pp. S586-S586 ◽  
Author(s):  
Kazuo Hashikawa ◽  
Hidefumi Yoshida ◽  
Nobukatsu Sawamoto ◽  
Shigetoshi Takaya ◽  
Chihiro Namiki ◽  
...  
Keyword(s):  
Parkinson’S Disease ◽  
Parkinson's Disease ◽  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Nicotinic Acetylcholine
Sign in / Sign up

Export Citation Format

Share Document