Electrostatic radius of the gramicidin channel determined from voltage dependence of H+ ion conductance

Respiration-dependent responses of a pH probe (fluorescein isothiocyanate, FITC), covalently bound to the membrane proteins of mitochondria and submitochondrial particles (SMP) have been studied. A spectral shift indicating FITC deprotonation was observed when respiration was activated in coupled mitochondria. Such a response was increased by valinomycin and reduced by uncoupler. Some FITC deprotonation was detected in the presence of excess of an uncoupler, but the response was smaller and insensitive to valinomycin. FITC deprotonation was also observed in submitochondrial particles after succinate addition. In this case it was not affected by uncoupler. Increase in the buffer concentration was found to (i) decrease the FITC response and (ii) increase the rate of uncoupled respiration in both mitochondria and submitochondrial particles. The results are consistent with the assumption that respiration initiates appearance of local H + activity gradients on the inner side of the internal mitochondrial membrane during the steady-state H + pumping. We suggest that the formation of this gradient is due to kinetic barrier to proton transfer from the bulk phase to the respiratory proton pump vicinity.z 1998 Federation of European Biochemical Societies.

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“…The presence of a similar barrier was reported earlier [16, 17]for H + transfer through the gramicidin channel inserted in BLM.…”

Section: Resultssupporting

confidence: 83%

Detection of the local H⁺ gradients on the internal mitochondrial membrane

Krasinskaya¹,

Lapin²,

Yaguzhinsky³

1998

FEBS Letters

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“…-82...-a! than the physical radius of the lumen, z2.1 A (Jordan, 1981(Jordan, , 1984aLevitt and Decker, 1988). As already pointed out, for the ionic strengths considered (0.01 to 2.5 M), results independent of system size are obtained with a radial cutoff of 31.25 A and an axial cutoff of ±95 A.…”

Section: S-mentioning

confidence: 66%

“…When it is narrow (e.g., gramicidin or K-channel), e is not known. The membrane width is W and the pore's electrical radius (a quantity somewhat larger than its physical radius: Jordan, 1981Jordan, , 1984aLevitt and Decker, 1988) is a,. In the equilibrium electrolyte (the regions to the right of z = L and the left of z = -L and, in the case of a nonselective channel, the channel itself), the Poisson-Boltzmann equation applies…”

Section: Introductionmentioning

confidence: 99%

How electrolyte shielding influences the electrical potential in transmembrane ion channels

Jordan

Bacquet

McCammon

et al. 1989

Biophysical Journal

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The electrical potential due to fixed charge distributions is strongly altered in the vicinity of a membrane and notably dependent on aqueous electrolyte concentration. We present an efficient way to solve the nonlinear Poisson-Boltzmann equation applicable to general cylindrically symmetric dielectric geometries. It generalizes Gouy-Chapman theory to systems containing transmembrane channels. The method is applied to three channel systems: gramicidin, gap junction, and porin. We find that for a long, narrow channel such as gramicidin concentration variation has little influence on the electrical image barrier to ion permeation. However, electrolyte shielding reduces the image induced contribution to the energy required for multiple occupancy. In addition, the presence of electrolyte significantly affects the voltage profile due to an applied potential, substantially compressing the electric field to the immediate vicinity of the pore itself. In the large diameter channels, where bulk electrolyte may be assumed to enter the pore, the electrolyte greatly reduces the image barrier to ion permeation. At physiological ionic strengths this barrier is negligible and the channel may be readily multiply occupied. At all ionic strengths considered (l greater than 0.005 M) the image barrier saturates rapidly and is essentially constant more than one channel radius from the entrance to the pore. At lower ionic strengths (l less than 0.016 M) there are noticeable (greater than 20 mV) energy penalties associated with multiple occupancy.

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“…2 As LqTx binding is inhibited by depolarization, gramicidin was used to increase the resting membrane potential and binding affinity. In the Na ϩ -free, choline-substituted isotonic solution used for binding assays, gramicidin allows K ϩ efflux without choline influx (40) to increase the negative membrane potential. Addition of gramicidin increased the receptor occupancy at 0.2 nM 1A).…”

Section: ␣-Scorpion Toxin Binding To Stably and Transiently Expressed Namentioning

confidence: 99%

Molecular Determinants of High Affinity Binding of α-Scorpion Toxin and Sea Anemone Toxin in the S3-S4 Extracellular Loop in Domain IV of the Na+ Channel α Subunit

Rogers

Tanada

et al. 1996

Journal of Biological Chemistry

415

517

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␣-Scorpion toxins and sea anemone toxins bind to a common extracellular site on the Na ؉ channel and inhibit fast inactivation. Basic amino acids of the toxins and domains I and IV of the Na ؉ channel ␣ subunit have been previously implicated in toxin binding. To identify acidic residues required for toxin binding, extracellular acidic amino acids in domains I and IV of the type IIa Na ؉ channel ␣ subunit were converted to neutral or basic amino acids using site-directed mutagenesis, and altered channels were transiently expressed in tsA-201 cells and tested for 125 I-␣-scorpion toxin binding. Conversion of Glu 1613 at the extracellular end of transmembrane segment IVS3 to Arg or His blocked measurable ␣-scorpion toxin binding, but did not affect the level of expression or saxitoxin binding affinity. Conversion of individual residues in the IVS3-S4 extracellular loop to differently charged residues or to Ala identified seven additional residues whose mutation caused significant effects on binding of ␣-scorpion toxin or sea anemone toxin. Moreover, chimeric Na ؉ channels in which amino acid residues at the extracellular end of segment IVS3 of the ␣ subunit of cardiac Na ؉ channels were substituted into the type IIa channel sequence had reduced affinity for ␣-scorpion toxin characteristic of cardiac Na ؉ channels. Electrophysiological analysis showed that E1613R has 62-and 82-fold lower affinities for ␣-scorpion and sea anemone toxins, respectively. Dissociation of ␣-scorpion toxin is substantially accelerated at all potentials compared to wild-type channels. ␣-Scorpion toxin binding to wild type and E1613R had similar voltage dependence, which was slightly more positive and steeper than the voltage dependence of steady-state inactivation. These results indicate that nonidentical amino acids of the IVS3-S4 loop participate in ␣-scorpion toxin and sea anemone toxin binding to overlapping sites and that neighboring amino acid residues in the IVS3 segment contribute to the difference in ␣-scorpion toxin binding affinity between cardiac and neuronal Na ؉ channels. The results also support the hypothesis that this region of the Na ؉ channel is important for coupling channel activation to fast inactivation.Voltage-gated Na ϩ channels are responsible for the conduction of electrical impulses in most excitable tissues (1). The importance of their function is demonstrated by the effects of Na ϩ channel-specific neurotoxins that bind to at least six different receptor sites on the Na ϩ channel molecule and disrupt its normal behavior (reviewed in Refs. 2 and 3). These natural toxins are powerful tools for understanding and correlating ion channel structure and function, as exemplified by identification of molecular determinants for binding of the pore blocker tetrodotoxin, which has provided important information about the structure of the ion selectivity filter and pore (3, 4). Similarly, the identification of molecular determinants for binding of toxins that modify activation or inactivation will likely provide important ...

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Electrostatic radius of the gramicidin channel determined from voltage dependence of H+ ion conductance

Cited by 30 publications

References 13 publications

Detection of the local H⁺ gradients on the internal mitochondrial membrane

Detection of the local H⁺ gradients on the internal mitochondrial membrane

How electrolyte shielding influences the electrical potential in transmembrane ion channels

Molecular Determinants of High Affinity Binding of α-Scorpion Toxin and Sea Anemone Toxin in the S3-S4 Extracellular Loop in Domain IV of the Na+ Channel α Subunit

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Electrostatic radius of the gramicidin channel determined from voltage dependence of H+ ion conductance

Cited by 30 publications

References 13 publications

Detection of the local H+ gradients on the internal mitochondrial membrane

Detection of the local H+ gradients on the internal mitochondrial membrane

How electrolyte shielding influences the electrical potential in transmembrane ion channels

Molecular Determinants of High Affinity Binding of α-Scorpion Toxin and Sea Anemone Toxin in the S3-S4 Extracellular Loop in Domain IV of the Na+ Channel α Subunit

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Product

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Detection of the local H⁺ gradients on the internal mitochondrial membrane

Detection of the local H⁺ gradients on the internal mitochondrial membrane