Click here to close
Hello! We notice that you are using Internet Explorer, which is not supported by Xenbase and may cause the site to display incorrectly.
We suggest using a current version of Chrome,
FireFox, or Safari.
Specific neosaxitoxin interactions with the Na+ channel outer vestibule determined by mutant cycle analysis.
Penzotti JL
,
Lipkind G
,
Fozzard HA
,
Dudley SC
.
???displayArticle.abstract???
The voltage-gated Na+ channel alpha-subunit consists of four homologous domains arranged circumferentially to form the pore. Several neurotoxins, including saxitoxin (STX), block the pore by binding to the outer vestibule of this permeation pathway, which is composed of four pore-forming loops (P-loops), one from each domain. Neosaxitoxin (neoSTX) is a variant of STX that differs only by having an additional hydroxyl group at the N1 position of the 1,2,3 guanidinium (N1-OH). We used this structural variant in mutant cycle experiments to determine interactions of the N1-OH and its guanidinium with the outer vestibule. NeoSTX had a higher affinity for the adult rat skeletal muscle Na+ channel (muI or Scn4a) than for STX (DeltaG approximately = 1.3 kcal/mol). Mutant cycle analysis identified groups that potentially interacted with each other. The N1 toxin site interacted most strongly with muI Asp-400 and Tyr-401. The interaction between the N1-OH of neoSTX and Tyr-401 was attractive (DeltaDeltaG = -1.3 +/- 0.1 kcal/mol), probably with formation of a hydrogen bond. A second possible attractive interaction to Asp-1532 was identified. There was repulsion between Asp-400 and the N1-OH (DeltaDeltaG = 1.4 +/- 0.1 kcal/mol), and kinetic analysis further suggested that the N1-OH was interacting negatively with Asp-400 at the transition state. Changes in pH altered the affinity of neoSTX, as would be expected if the N1-OH site were partially deprotonated. These interactions offer an explanation for most of the difference in blocking efficacy between neoSTX and STX and for the sensitivity of neoSTX to pH. Kinetic analysis suggested significant differences in coupling energies between the transition and the equilibrium, bound states. This is the first report to identify points of interaction between a channel and a non-peptide toxin. This interaction pattern was consistent with previous proposals describing the interactions of STX with the outer vestibule (Lipkind, G. M., and H. A. Fozzard. 1994. Biophys. J. 66:1-13; Penzotti, J. L., G. Lipkind, H. A. Fozzard, and S. C. Dudley, Jr. 1998. Biophys. J. 75:2647-2657).
Braiman,
A large photolysis-induced pKa increase of the chromophore counterion in bacteriorhodopsin: implications for ion transport mechanisms of retinal proteins.
1996, Pubmed
Braiman,
A large photolysis-induced pKa increase of the chromophore counterion in bacteriorhodopsin: implications for ion transport mechanisms of retinal proteins.
1996,
Pubmed
Chang,
Predominant interactions between mu-conotoxin Arg-13 and the skeletal muscle Na+ channel localized by mutant cycle analysis.
1998,
Pubmed
,
Xenbase
Chen,
Cysteine mapping in the ion selectivity and toxin binding region of the cardiac Na+ channel pore.
1997,
Pubmed
Chen,
Aspartate substitutions establish the concerted action of P-region glutamates in repeats I and III in forming the protonation site of L-type Ca2+ channels.
1997,
Pubmed
,
Xenbase
Chiamvimonvat,
Depth asymmetries of the pore-lining segments of the Na+ channel revealed by cysteine mutagenesis.
1996,
Pubmed
,
Xenbase
Davoodi,
Abnormally high pKa of an active-site glutamic acid residue in Bacillus circulans xylanase. The role of electrostatic interactions.
1995,
Pubmed
Favre,
On the structural basis for ionic selectivity among Na+, K+, and Ca2+ in the voltage-gated sodium channel.
1996,
Pubmed
,
Xenbase
Favre,
Specificity for block by saxitoxin and divalent cations at a residue which determines sensitivity of sodium channel subtypes to guanidinium toxins.
1995,
Pubmed
,
Xenbase
Fersht,
The folding of an enzyme. I. Theory of protein engineering analysis of stability and pathway of protein folding.
1992,
Pubmed
Guo,
Kinetic basis for insensitivity to tetrodotoxin and saxitoxin in sodium channels of canine heart and denervated rat skeletal muscle.
1987,
Pubmed
Hidalgo,
Revealing the architecture of a K+ channel pore through mutant cycles with a peptide inhibitor.
1995,
Pubmed
,
Xenbase
Hille,
The receptor for tetrodotoxin and saxitoxin. A structural hypothesis.
1975,
Pubmed
Hu,
Interactions of neosaxitoxin with the sodium channel of the frog skeletal muscle fiber.
1991,
Pubmed
Kao,
The active guanidinium group of saxitoxin and neosaxitoxin identified by the effects of pH on their activities on squid axon.
1983,
Pubmed
Kao,
Active groups of saxitoxin and tetrodotoxin as deduced from actions of saxitoxin analogues on frog muscle and squid axon.
1982,
Pubmed
Kao,
Structure-activity relations of tetrodotoxin, saxitoxin, and analogues.
1986,
Pubmed
Kirsch,
Differential effects of sulfhydryl reagents on saxitoxin and tetrodotoxin block of voltage-dependent Na channels.
1994,
Pubmed
Kontis,
Site-directed mutagenesis of the putative pore region of the rat IIA sodium channel.
1993,
Pubmed
,
Xenbase
Langsetmo,
The conserved, buried aspartic acid in oxidized Escherichia coli thioredoxin has a pKa of 7.5. Its titration produces a related shift in global stability.
1991,
Pubmed
Levitt,
Aromatic rings act as hydrogen bond acceptors.
1988,
Pubmed
Lipkind,
A structural model of the tetrodotoxin and saxitoxin binding site of the Na+ channel.
1994,
Pubmed
Matouschek,
Mapping the transition state and pathway of protein folding by protein engineering.
1989,
Pubmed
Moczydlowski,
Voltage-dependent blockade of muscle Na+ channels by guanidinium toxins.
1984,
Pubmed
Noda,
A single point mutation confers tetrodotoxin and saxitoxin insensitivity on the sodium channel II.
1989,
Pubmed
,
Xenbase
Penzotti,
Differences in saxitoxin and tetrodotoxin binding revealed by mutagenesis of the Na+ channel outer vestibule.
1998,
Pubmed
,
Xenbase
Pérez-García,
Structure of the sodium channel pore revealed by serial cysteine mutagenesis.
1996,
Pubmed
,
Xenbase
Rousso,
pKa of the protonated Schiff base and aspartic 85 in the bacteriorhodopsin binding site is controlled by a specific geometry between the two residues.
1995,
Pubmed
Sampogna,
Environmental effects on the protonation states of active site residues in bacteriorhodopsin.
1994,
Pubmed
Schantz,
Letter: The structure of saxitoxin.
1975,
Pubmed
Schreiber,
Energetics of protein-protein interactions: analysis of the barnase-barstar interface by single mutations and double mutant cycles.
1995,
Pubmed
Shimizu,
Presence of four toxins in red tide infested clams and cultured Gonyaulax tamarensis cells.
1975,
Pubmed
Shimizu,
Chemistry and biochemistry of saxitoxin analogues and tetrodotoxin.
1986,
Pubmed
Strichartz,
Structural determinants of the affinity of saxitoxin for neuronal sodium channels. Electrophysiological studies on frog peripheral nerve.
1984,
Pubmed
Sun,
On the structural basis for size-selective permeation of organic cations through the voltage-gated sodium channel. Effect of alanine mutations at the DEKA locus on selectivity, inhibition by Ca2+ and H+, and molecular sieving.
1997,
Pubmed
Terlau,
Mapping the site of block by tetrodotoxin and saxitoxin of sodium channel II.
1991,
Pubmed
,
Xenbase
Woodhull,
Ionic blockage of sodium channels in nerve.
1973,
Pubmed
Yamagishi,
Topology of the P segments in the sodium channel pore revealed by cysteine mutagenesis.
1997,
Pubmed
Yang,
Actions of decarbamoyloxysaxitoxin and decarbamoylneosaxitoxin on the frog skeletal muscle fiber.
1992,
Pubmed
Zhang,
Effects of external protons on single cardiac sodium channels from guinea pig ventricular myocytes.
1991,
Pubmed
