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Cystic fibrosis transmembrane conductance regulator. Physical basis for lyotropic anion selectivity patterns.

Smith SS, Steinle ED, Meyerhoff ME, Dawson DC - J. Gen. Physiol. (1999)

Bottom Line: The calculated energies of anion-channel interaction, derived from measurements of either permeability or binding, varied as a linear function of inverse ionic radius (1/r), as expected from a Born-type model of ion charging in a medium characterized by an effective dielectric constant of 19.These large anions also bind more tightly for the same reason: the reduced energy of hydration allows the net transfer energy (the well depth) to be more negative.Anions that are smaller (more difficult to dehydrate) than Cl are energetically retarded from entering the channel, while the larger (more readily dehydrated) anions are retarded in their passage by "sticking" within the channel.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, University of Michigan, Ann Arbor, Michigan 48109, USA.

ABSTRACT
The cystic fibrosis transmembrane conductance regulator (CFTR) Cl channel exhibits lyotropic anion selectivity. Anions that are more readily dehydrated than Cl exhibit permeability ratios (P(S)/P(Cl)) greater than unity and also bind more tightly in the channel. We compared the selectivity of CFTR to that of a synthetic anion-selective membrane [poly(vinyl chloride)-tridodecylmethylammonium chloride; PVC-TDMAC] for which the nature of the physical process that governs the anion-selective response is more readily apparent. The permeability and binding selectivity patterns of CFTR differed only by a multiplicative constant from that of the PVC-TDMAC membrane; and a continuum electrostatic model suggested that both patterns could be understood in terms of the differences in the relative stabilization of anions by water and the polarizable interior of the channel or synthetic membrane. The calculated energies of anion-channel interaction, derived from measurements of either permeability or binding, varied as a linear function of inverse ionic radius (1/r), as expected from a Born-type model of ion charging in a medium characterized by an effective dielectric constant of 19. The model predicts that large anions, like SCN, although they experience weaker interactions (relative to Cl) with water and also with the channel, are more permeant than Cl because anion-water energy is a steeper function of 1/r than is the anion-channel energy. These large anions also bind more tightly for the same reason: the reduced energy of hydration allows the net transfer energy (the well depth) to be more negative. This simple selectivity mechanism that governs permeability and binding acts to optimize the function of CFTR as a Cl filter. Anions that are smaller (more difficult to dehydrate) than Cl are energetically retarded from entering the channel, while the larger (more readily dehydrated) anions are retarded in their passage by "sticking" within the channel.

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Iodide block of CFTR current. (A) The solid line is the cAMP-activated current (10 μM forskolin + 1 mM IBMX) in standard frog ringer (materials and methods) for an oocyte expressing wtCFTR. The dashed line is the current 2 min after the addition of 5 mM NaI to the perfusate. (B) The solid line is the cAMP-activated current (10 μM forskolin + 1 mM IBMX) in a reduced Cl frog ringer (30 mM Cl; 70 mM aspartate) for an oocyte expressing wtCFTR. The dashed line is the current 2 min after the addition of 5 mM NaI to the perfusate. The data was collected using a ramp protocol as noted in materials and methods.
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Figure 7: Iodide block of CFTR current. (A) The solid line is the cAMP-activated current (10 μM forskolin + 1 mM IBMX) in standard frog ringer (materials and methods) for an oocyte expressing wtCFTR. The dashed line is the current 2 min after the addition of 5 mM NaI to the perfusate. (B) The solid line is the cAMP-activated current (10 μM forskolin + 1 mM IBMX) in a reduced Cl frog ringer (30 mM Cl; 70 mM aspartate) for an oocyte expressing wtCFTR. The dashed line is the current 2 min after the addition of 5 mM NaI to the perfusate. The data was collected using a ramp protocol as noted in materials and methods.

Mentions: In terms of permeability selectivity, iodide stands out as an anomaly. In Fig. 2 A, it can be seen that the value of PI/PCl determined for human wtCFTR expressed in Xenopus oocytes is well below that predicted for an ion that is easier to dehydrate than Cl. Tabcharani et al. 1997 provided evidence that CFTR can exhibit a higher value of PI/PCl (2.1; Fig. 2 A, asterisk), but that the ion causes a rapid modification of CFTR that leads to the lower value of 0.2–0.4 that is most often reported (Anderson et al. 1991; Sheppard et al. 1993; Mansoura et al. 1998). The higher value of PI/PCl would place iodide in its predicted position on the 1/r plot (Fig. 2 A). Using human wtCFTR expressed in Xenopus oocytes, we have not detected any evidence of this higher PI/PCl using a panel of various mole fractions of I:Cl, as well as various voltage clamping protocols (data not shown). It may be, however, that the conversion is simply too fast to resolve in this experimental setting. In accord with the hypothesis of Tabcharani et al. 1997 that I is inducing a modification of CFTR, we have observed a block of CFTR by I that is qualitatively distinct from the block seen with the other permeant ions (see Mansoura et al. 1998). Block by a small amount of I in the external bath is weakly voltage dependent in the negative quadrant, as shown in Fig. 7, whereas the block seen with most permeant ions is largely voltage independent (Mansoura et al. 1998). The efficacy of block is enhanced if the external concentration of Cl is lowered (by substitution with aspartate), as shown in Fig. 7 B, suggesting that Cl and I ions are competing for the same site(s). Iodide has a tendency to form interhalogens, particularly triiodide, a reactive species known to attack cysteine, tyrosine, and histidine residues within proteins, but the addition of sodium thiosulfate, which reduces I3, did not alter the observed effects. Furthermore, increasing concentrations of I3 (made by adding I2 to NaI solution) appeared to be quite toxic above 500 μM I3, leading to uncontrollable increases in oocyte conductance. While in solutions near or below 100 μM, the I3 appeared to be unstable in our standard amphibian Ringer's (as judged colorimetrically by clarification of the yellowish-red color of the I3), it was possible to make a fresh I3 solution and feel relatively confident that the oocyte was seeing 10–100 μM I3, in which case the effect on CFTR was an irreversible, voltage-independent block. However, we cannot rule out the possibility that I3 may be forming within the lumen, leading to a mild chemical modification of CFTR. Data from mutagenesis experiments support the view that iodide permeation is most properly thought of as a special case in that mutations that affect PI/PCl do not appear to dramatically alter the permeability ratios for other permeant ions (Anderson et al. 1991; Tabcharani et al. 1997; Mansoura et al. 1998).


Cystic fibrosis transmembrane conductance regulator. Physical basis for lyotropic anion selectivity patterns.

Smith SS, Steinle ED, Meyerhoff ME, Dawson DC - J. Gen. Physiol. (1999)

Iodide block of CFTR current. (A) The solid line is the cAMP-activated current (10 μM forskolin + 1 mM IBMX) in standard frog ringer (materials and methods) for an oocyte expressing wtCFTR. The dashed line is the current 2 min after the addition of 5 mM NaI to the perfusate. (B) The solid line is the cAMP-activated current (10 μM forskolin + 1 mM IBMX) in a reduced Cl frog ringer (30 mM Cl; 70 mM aspartate) for an oocyte expressing wtCFTR. The dashed line is the current 2 min after the addition of 5 mM NaI to the perfusate. The data was collected using a ramp protocol as noted in materials and methods.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2230651&req=5

Figure 7: Iodide block of CFTR current. (A) The solid line is the cAMP-activated current (10 μM forskolin + 1 mM IBMX) in standard frog ringer (materials and methods) for an oocyte expressing wtCFTR. The dashed line is the current 2 min after the addition of 5 mM NaI to the perfusate. (B) The solid line is the cAMP-activated current (10 μM forskolin + 1 mM IBMX) in a reduced Cl frog ringer (30 mM Cl; 70 mM aspartate) for an oocyte expressing wtCFTR. The dashed line is the current 2 min after the addition of 5 mM NaI to the perfusate. The data was collected using a ramp protocol as noted in materials and methods.
Mentions: In terms of permeability selectivity, iodide stands out as an anomaly. In Fig. 2 A, it can be seen that the value of PI/PCl determined for human wtCFTR expressed in Xenopus oocytes is well below that predicted for an ion that is easier to dehydrate than Cl. Tabcharani et al. 1997 provided evidence that CFTR can exhibit a higher value of PI/PCl (2.1; Fig. 2 A, asterisk), but that the ion causes a rapid modification of CFTR that leads to the lower value of 0.2–0.4 that is most often reported (Anderson et al. 1991; Sheppard et al. 1993; Mansoura et al. 1998). The higher value of PI/PCl would place iodide in its predicted position on the 1/r plot (Fig. 2 A). Using human wtCFTR expressed in Xenopus oocytes, we have not detected any evidence of this higher PI/PCl using a panel of various mole fractions of I:Cl, as well as various voltage clamping protocols (data not shown). It may be, however, that the conversion is simply too fast to resolve in this experimental setting. In accord with the hypothesis of Tabcharani et al. 1997 that I is inducing a modification of CFTR, we have observed a block of CFTR by I that is qualitatively distinct from the block seen with the other permeant ions (see Mansoura et al. 1998). Block by a small amount of I in the external bath is weakly voltage dependent in the negative quadrant, as shown in Fig. 7, whereas the block seen with most permeant ions is largely voltage independent (Mansoura et al. 1998). The efficacy of block is enhanced if the external concentration of Cl is lowered (by substitution with aspartate), as shown in Fig. 7 B, suggesting that Cl and I ions are competing for the same site(s). Iodide has a tendency to form interhalogens, particularly triiodide, a reactive species known to attack cysteine, tyrosine, and histidine residues within proteins, but the addition of sodium thiosulfate, which reduces I3, did not alter the observed effects. Furthermore, increasing concentrations of I3 (made by adding I2 to NaI solution) appeared to be quite toxic above 500 μM I3, leading to uncontrollable increases in oocyte conductance. While in solutions near or below 100 μM, the I3 appeared to be unstable in our standard amphibian Ringer's (as judged colorimetrically by clarification of the yellowish-red color of the I3), it was possible to make a fresh I3 solution and feel relatively confident that the oocyte was seeing 10–100 μM I3, in which case the effect on CFTR was an irreversible, voltage-independent block. However, we cannot rule out the possibility that I3 may be forming within the lumen, leading to a mild chemical modification of CFTR. Data from mutagenesis experiments support the view that iodide permeation is most properly thought of as a special case in that mutations that affect PI/PCl do not appear to dramatically alter the permeability ratios for other permeant ions (Anderson et al. 1991; Tabcharani et al. 1997; Mansoura et al. 1998).

Bottom Line: The calculated energies of anion-channel interaction, derived from measurements of either permeability or binding, varied as a linear function of inverse ionic radius (1/r), as expected from a Born-type model of ion charging in a medium characterized by an effective dielectric constant of 19.These large anions also bind more tightly for the same reason: the reduced energy of hydration allows the net transfer energy (the well depth) to be more negative.Anions that are smaller (more difficult to dehydrate) than Cl are energetically retarded from entering the channel, while the larger (more readily dehydrated) anions are retarded in their passage by "sticking" within the channel.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, University of Michigan, Ann Arbor, Michigan 48109, USA.

ABSTRACT
The cystic fibrosis transmembrane conductance regulator (CFTR) Cl channel exhibits lyotropic anion selectivity. Anions that are more readily dehydrated than Cl exhibit permeability ratios (P(S)/P(Cl)) greater than unity and also bind more tightly in the channel. We compared the selectivity of CFTR to that of a synthetic anion-selective membrane [poly(vinyl chloride)-tridodecylmethylammonium chloride; PVC-TDMAC] for which the nature of the physical process that governs the anion-selective response is more readily apparent. The permeability and binding selectivity patterns of CFTR differed only by a multiplicative constant from that of the PVC-TDMAC membrane; and a continuum electrostatic model suggested that both patterns could be understood in terms of the differences in the relative stabilization of anions by water and the polarizable interior of the channel or synthetic membrane. The calculated energies of anion-channel interaction, derived from measurements of either permeability or binding, varied as a linear function of inverse ionic radius (1/r), as expected from a Born-type model of ion charging in a medium characterized by an effective dielectric constant of 19. The model predicts that large anions, like SCN, although they experience weaker interactions (relative to Cl) with water and also with the channel, are more permeant than Cl because anion-water energy is a steeper function of 1/r than is the anion-channel energy. These large anions also bind more tightly for the same reason: the reduced energy of hydration allows the net transfer energy (the well depth) to be more negative. This simple selectivity mechanism that governs permeability and binding acts to optimize the function of CFTR as a Cl filter. Anions that are smaller (more difficult to dehydrate) than Cl are energetically retarded from entering the channel, while the larger (more readily dehydrated) anions are retarded in their passage by "sticking" within the channel.

Show MeSH
Related in: MedlinePlus