NMR structure determination of α-conotoxin BuIA, a novel neuronal nicotinic acetylcholine receptor antagonist with an unusual 4/4 disulfide scaffold (original) (raw)
Related papers
Journal of Biological Chemistry, 2003
We report the solution three-dimensional structure of an ␣A-conotoxin EIVA determined by nuclear magnetic resonance spectroscopy and restrained molecular dynamics. The ␣A-conotoxin EIVA consists of 30 amino acids representing the largest peptide among the ␣/␣Afamily conotoxins discovered so far and targets the neuromuscular nicotinic acetylcholine receptor with high affinity. ␣A-Conotoxin EIVA consists of three distinct structural domains. The first domain is mainly composed of the Cys 3-Cys 11-disulfide loop and is structurally ill-defined with a large backbone root mean square deviation of 1.91 Å. The second domain formed by residues His 12-Hyp 21 is extremely well defined with a backbone root mean square deviation of 0.52 Å, thus forming a sturdy stem for the entire molecule. The third C-terminal domain formed by residues Hyp 22-Gly 29 shows an intermediate structural order having a backbone root mean square deviation of 1.04 Å. A structurally ill-defined N-terminal first loop domain connected to a rigid central molecular stem seems to be the general structural feature of the ␣A-conotoxin subfamily. A detailed structural comparison between ␣A-conotoxin EIVA and ␣A-conotoxin PIVA suggests that the higher receptor affinity of ␣A-conotoxin EIVA than ␣A-conotoxin PIVA might originate from different steric disposition and charge distribution in the second loop "handle" motif.
Biochemical and Biophysical Research Communications, 2005
a-Conotoxin PIA is a novel nicotinic acetylcholine receptor (nAChR) antagonist isolated from Conus purpurascens that targets nAChR subtypes containing a6 and a3 subunits. a-conotoxin PIA displays 75-fold higher affinity for rat a6/a3b2b3 nAChRs than for rat a3b2 nAChRs. We have determined the three-dimensional structure of a-conotoxin PIA by nuclear magnetic resonance spectroscopy. The a-conotoxin PIA has an ''x-shaped'' overall topology as other a4/7 subfamily conotoxins. Yet, unlike other neuronally targeted a4/7-conotoxins, its N-terminal tail Arg 1-Asp 2-Pro 3 protrudes out of its main molecular body because Asp 2-Pro 3-Cys 4-Cys 5 forms a stable type I b-turn. In addition, a kink introduced by Pro 15 in the second loop of this toxin provides a distinct steric and electrostatic environment from those in a-conotoxins MII and GIC. By comparing the structure of a-conotoxin PIA with other functionally related a-conotoxins we suggest structural features in a-conotoxin PIA that may be associated with its unique receptor recognition profile.
Biochemical and Biophysical Research Communications, 2006
a-Conotoxin OmIA from Conus omaria is the only a-conotoxin that shows a $20-fold higher affinity to the a3b2 over the a6b2 subtype of nicotinic acetylcholine receptor. We have determined a three-dimensional structure of a-conotoxin OmIA by nuclear magnetic resonance spectroscopy. a-Conotoxin OmIA has an ''x-shaped'' overall topology with His 5-Asn 12 forming an a-helix. Structural features of a-conotoxin OmIA responsible for its selectivity are suggested by comparing its surface characteristics with other functionally related a4/7 subfamily conotoxins. Reduced size of the hydrophilic area in a-conotoxin OmIA seems to be associated with the reduced affinity towards the a6b2 nAChR subtype.
Structure, 1996
Background: ␣-Conotoxins are peptide toxins, isolated from Conus snails, that block the nicotinic acetylcholine receptor (nAChR). The 16-residue peptides PnIA and PnIB from Conus pennaceus incorporate the same disulfide framework as other ␣-conotoxins but differ in function from most ␣-conotoxins by blocking the neuronal nAChR, rather than the skeletal muscle subtype. The crystal structure determination of PnIA was undertaken to identify structural and surface features that might be important for biological activity.
Biochemistry
We report the purification and characterization of a new conotoxin from the venom of Conus radiatus. The peptide, RS-conotoxin RVIIIA (RS-RVIIIA), is biochemically unique with respect to its amino acid sequence, post-translational modification, and molecular targets. In comparison to other nicotinic antagonists from Conus venoms, RS-RVIIIA exhibits an unusually broad targeting specificity for nicotinic acetylcholine receptor (nAChR) subtypes, as assayed by electrophysiology. The toxin is paralytic to mice and fish, consistent with its nearly irreversible block of the neuromuscular nAChR. Similar to other antagonists of certain neuronal nAChRs, the toxin also elicits seizures in mice upon intracranial injection. The only previously characterized conotoxin from the S superfamily, σ-conotoxin GVIIIA, is a specific competitive antagonist of the 5-HT 3 receptor; thus, RS-RVIIIA defines a novel family of nicotinic antagonists within the S superfamily. All previously characterized competitive conotoxin nAChR antagonists have been members of the A superfamily of conotoxins. Our working hypothesis is that the particular group of fish-hunting Conus species that includes Conus radiatus uses the RS-conotoxin family to target the muscle nAChR and paralyze prey.
Marine Drugs, 2015
In the present study, we synthesized and, structurally and functionally characterized a novel α4/7-conotoxin Mr1.7 (PECCTHPACHVSHPELC-NH2), which was previously identified by cDNA libraries from Conus marmoreus in our lab. The NMR solution structure showed that Mr1.7 contained a 310-helix from residues Pro7 to His10 and a type I β-turn from residues Pro14 to Cys17. Electrophysiological results showed that Mr1.7 selectively inhibited the α3β2, α9α10 and α6/α3β2β3 neuronal nicotinic acetylcholine receptors (nAChRs) with an IC50 of 53.1 nM, 185.7 nM and 284.2 nM, respectively, but showed no inhibitory activity on other nAChR subtypes. Further structure-activity studies of Mr1.7 demonstrated that the PE residues at the N-terminal sequence of Mr1.7 were important for modulating its selectivity, and the replacement of Glu2 by Ala resulted in a significant increase in potency and selectivity to the α3β2 nAChR. Furthermore, the substitution of Ser12 with Asn in the loop2 significantly incre...
Biochemistry, 1999
The three-dimensional solution conformation of an 11-residue antitoxic analogue of R-conotoxin GI, des-Glu1-[Cys3Ala]-des-Cys13-conotoxin GI (CANPACGRHYS-NH 2 , designated "GI-15" henceforth), has been determined using two-dimensional 1 H NMR spectroscopy. The disulfide loop region (1C-6C) and the C-terminal tail (8R-11S) are connected by a flexible hinge formed near 7G, and the pairwise backbone rmsds for the former and the latter are 0.58 and 0.65 Å, respectively. Superpositioning GI-15 with the structure of R-conotoxin GI shows that the two share an essentially identical fold in the common first disulfide loop region (1C-6C). However, the absence of the second disulfide loop in GI-15 results in segmental motion of the C-terminal half, causing the key receptor subtype selectivity residue 8R (Arg9 in R-conotoxin GI) to lose its native spatial orientation. The combined features of structural equivalence in the disulfide loop and a mobile C-terminal tail appear to be responsible for the activity of GI-15 as a competitive antagonist against native toxin. Electrostatic surface potential comparisons of the first disulfide region of GI-15 with other R-conotoxins or receptor-bound states of acetylcholine and d-tubocurarine show a common protruding surface that may serve as the minimal binding determinant for the neuromuscular acetylcholine receptor R 1 -subunit. On the basis of the original "Conus toxin macrosite model" [Olivera, B. FIGURE 3: Stereoviews of the 30 final structures of GI-15 superimposed over the backbone atoms (N, C R , C′, and O) of (a) residues 1-6 and (b) residues 7-11. Residues 1-6 are not shown in panel b. Selected residues are labeled.
A New α-Conotoxin Which Targets α3β2 Nicotinic Acetylcholine Receptors
Journal of Biological Chemistry, 1996
We have isolated a 16-amino acid peptide from the venom of the marine snail Conus magus which potently blocks nicotinic acetylcholine receptors (nAChRs) composed of ␣32 subunits. This peptide, named ␣-conotoxin MII, was identified by electrophysiologically screening venom fractions against cloned nicotinic receptors expressed in Xenopus oocytes. The peptide's structure, which has been confirmed by mass spectrometry and total chemical synthesis, differs significantly from those of all previously isolated ␣-conotoxins. Disulfide bridging, however, is conserved. The toxin blocks the response to acetylcholine in oocytes expressing ␣32 nAChRs with an IC 50 of 0.5 nM and is 2-4 orders of magnitude less potent on other nAChR subunit combinations. We have recently reported the isolation and characterization of ␣-conotoxin ImI, which selectively targets homomeric ␣7 neuronal nAChRs. Yet other ␣-conotoxins selectively block the muscle subtype of nAChR. Thus, it is increasingly apparent that ␣-conotoxins represent a significant resource for ligands with which to probe structure-function relationships of various nAChR subtypes.
Biochemistry
The three-dimensional solution conformation of an 11-residue antitoxic analogue of alpha-conotoxin GI, des-Glu1-[Cys3Ala]-des-Cys13-conotoxin GI (CANPACGRHYS-NH(2), designated "GI-15" henceforth), has been determined using two-dimensional (1)H NMR spectroscopy. The disulfide loop region (1C-6C) and the C-terminal tail (8R-11S) are connected by a flexible hinge formed near 7G, and the pairwise backbone rmsds for the former and the latter are 0.58 and 0.65 A, respectively. Superpositioning GI-15 with the structure of alpha-conotoxin GI shows that the two share an essentially identical fold in the common first disulfide loop region (1C-6C). However, the absence of the second disulfide loop in GI-15 results in segmental motion of the C-terminal half, causing the key receptor subtype selectivity residue 8R (Arg9 in alpha-conotoxin GI) to lose its native spatial orientation. The combined features of structural equivalence in the disulfide loop and a mobile C-terminal tail appear...