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Functional consequences of a decreased potassium affinity in a potassium channel pore. Ion interactions and C-type inactivation.

Ogielska EM, Aldrich RW - J. Gen. Physiol. (1999)

Bottom Line: However, we have found that this mutation also decreases the C-type inactivation rate of the channel.Our studies indicate that the C-type inactivation effects observed with substitutions at position A463 most likely result from changes in the pore occupancy of the channel, rather than a change in the C-type inactivation conformational change.We have found that a decrease in the potassium affinity of the internal ion binding site in the pore results in lowered (electrostatic) interactions among ions in the pore and as a result prolongs the time an ion remains bound at the external C-type inactivation site.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cellular Physiology, Howard Hughes Medical Institute, Stanford University, Stanford, California 94305, USA.

ABSTRACT
Ions bound near the external mouth of the potassium channel pore impede the C-type inactivation conformational change (Lopez-Barneo, J., T. Hoshi, S. Heinemann, and R. Aldrich. 1993. Receptors Channels. 1:61- 71; Baukrowitz, T., and G. Yellen. 1995. Neuron. 15:951-960). In this study, we present evidence that the occupancy of the C-type inactivation modulatory site by permeant ions is not solely dependent on its intrinsic affinity, but is also a function of the relative affinities of the neighboring sites in the potassium channel pore. The A463C mutation in the S6 region of Shaker decreases the affinity of an internal ion binding site in the pore (Ogielska, E.M., and R.W. Aldrich, 1998). However, we have found that this mutation also decreases the C-type inactivation rate of the channel. Our studies indicate that the C-type inactivation effects observed with substitutions at position A463 most likely result from changes in the pore occupancy of the channel, rather than a change in the C-type inactivation conformational change. We have found that a decrease in the potassium affinity of the internal ion binding site in the pore results in lowered (electrostatic) interactions among ions in the pore and as a result prolongs the time an ion remains bound at the external C-type inactivation site. We also present evidence that the C-type inactivation constriction is quite local and does not involve a general collapse of the selectivity filter. Our data indicate that in A463C potassium can bind within the selectivity filter without interfering with the process of C-type inactivation.

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State diagram model of external potassium block of  sodium currents in A463C. Sodium ions are represented by ○,  while potassium ions are shown as •. (Outlined scheme) In the  presence of external NMG+, external potassium first binds to the  C-type inactivation site but proceeds to a deeper, higher affinity  site in the pore. The occupancy of the deeper site by potassium  does not interfere with the onset of C-type inactivation. (Shaded  scheme) In the presence of external Na+, external potassium must  first displace the bound sodium ion before it can proceed to the  deeper site. After the higher affinity site is reached, the external  (C-type) site is refilled by a sodium ion.
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Figure 7: State diagram model of external potassium block of sodium currents in A463C. Sodium ions are represented by ○, while potassium ions are shown as •. (Outlined scheme) In the presence of external NMG+, external potassium first binds to the C-type inactivation site but proceeds to a deeper, higher affinity site in the pore. The occupancy of the deeper site by potassium does not interfere with the onset of C-type inactivation. (Shaded scheme) In the presence of external Na+, external potassium must first displace the bound sodium ion before it can proceed to the deeper site. After the higher affinity site is reached, the external (C-type) site is refilled by a sodium ion.

Mentions: Given the sensitivity of C-type inactivation to the presence of external ions (Lopez-Barneo et al., 1993; Baukrowitz and Yellen, 1995), the observation that it proceeds through a localized constriction of the outer mouth of the pore (Yellen et al., 1994; Liu et al., 1996), and the visualization of an ion bound at an external site in the crystal structure (Doyle et al., 1998), it is most likely that the site that controls C-type inactivation is the external site. Since the rate of C-type inactivation is unaffected when externally applied potassium is bound at its blocking site (Fig. 6 C), the high affinity site cannot be the external (C-type) site. External potassium presumably first binds to the C-type site but rapidly proceeds to a higher affinity blocking site, the occupancy of which does not interfere with C-type inactivation (Fig. 7, outlined scheme). The finding that external potassium still blocks sodium currents with a high affinity in the presence of external sodium indicates that the blocking ion presumably first displaces the bound sodium and subsequently binds at a higher affinity site in the pore. For example, sodium may bind and unbind rapidly at the C-type inactivation site while potassium enters deeper into the pore of the channel (Fig. 7, gray scheme). The rapid equilibration of the external (C-type) site is in agreement with the finding by Harris et al. (1998) that ions bound at the outermost site in the Shaker channel are in rapid equilibrium with the external solution.


Functional consequences of a decreased potassium affinity in a potassium channel pore. Ion interactions and C-type inactivation.

Ogielska EM, Aldrich RW - J. Gen. Physiol. (1999)

State diagram model of external potassium block of  sodium currents in A463C. Sodium ions are represented by ○,  while potassium ions are shown as •. (Outlined scheme) In the  presence of external NMG+, external potassium first binds to the  C-type inactivation site but proceeds to a deeper, higher affinity  site in the pore. The occupancy of the deeper site by potassium  does not interfere with the onset of C-type inactivation. (Shaded  scheme) In the presence of external Na+, external potassium must  first displace the bound sodium ion before it can proceed to the  deeper site. After the higher affinity site is reached, the external  (C-type) site is refilled by a sodium ion.
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Related In: Results  -  Collection

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Figure 7: State diagram model of external potassium block of sodium currents in A463C. Sodium ions are represented by ○, while potassium ions are shown as •. (Outlined scheme) In the presence of external NMG+, external potassium first binds to the C-type inactivation site but proceeds to a deeper, higher affinity site in the pore. The occupancy of the deeper site by potassium does not interfere with the onset of C-type inactivation. (Shaded scheme) In the presence of external Na+, external potassium must first displace the bound sodium ion before it can proceed to the deeper site. After the higher affinity site is reached, the external (C-type) site is refilled by a sodium ion.
Mentions: Given the sensitivity of C-type inactivation to the presence of external ions (Lopez-Barneo et al., 1993; Baukrowitz and Yellen, 1995), the observation that it proceeds through a localized constriction of the outer mouth of the pore (Yellen et al., 1994; Liu et al., 1996), and the visualization of an ion bound at an external site in the crystal structure (Doyle et al., 1998), it is most likely that the site that controls C-type inactivation is the external site. Since the rate of C-type inactivation is unaffected when externally applied potassium is bound at its blocking site (Fig. 6 C), the high affinity site cannot be the external (C-type) site. External potassium presumably first binds to the C-type site but rapidly proceeds to a higher affinity blocking site, the occupancy of which does not interfere with C-type inactivation (Fig. 7, outlined scheme). The finding that external potassium still blocks sodium currents with a high affinity in the presence of external sodium indicates that the blocking ion presumably first displaces the bound sodium and subsequently binds at a higher affinity site in the pore. For example, sodium may bind and unbind rapidly at the C-type inactivation site while potassium enters deeper into the pore of the channel (Fig. 7, gray scheme). The rapid equilibration of the external (C-type) site is in agreement with the finding by Harris et al. (1998) that ions bound at the outermost site in the Shaker channel are in rapid equilibrium with the external solution.

Bottom Line: However, we have found that this mutation also decreases the C-type inactivation rate of the channel.Our studies indicate that the C-type inactivation effects observed with substitutions at position A463 most likely result from changes in the pore occupancy of the channel, rather than a change in the C-type inactivation conformational change.We have found that a decrease in the potassium affinity of the internal ion binding site in the pore results in lowered (electrostatic) interactions among ions in the pore and as a result prolongs the time an ion remains bound at the external C-type inactivation site.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cellular Physiology, Howard Hughes Medical Institute, Stanford University, Stanford, California 94305, USA.

ABSTRACT
Ions bound near the external mouth of the potassium channel pore impede the C-type inactivation conformational change (Lopez-Barneo, J., T. Hoshi, S. Heinemann, and R. Aldrich. 1993. Receptors Channels. 1:61- 71; Baukrowitz, T., and G. Yellen. 1995. Neuron. 15:951-960). In this study, we present evidence that the occupancy of the C-type inactivation modulatory site by permeant ions is not solely dependent on its intrinsic affinity, but is also a function of the relative affinities of the neighboring sites in the potassium channel pore. The A463C mutation in the S6 region of Shaker decreases the affinity of an internal ion binding site in the pore (Ogielska, E.M., and R.W. Aldrich, 1998). However, we have found that this mutation also decreases the C-type inactivation rate of the channel. Our studies indicate that the C-type inactivation effects observed with substitutions at position A463 most likely result from changes in the pore occupancy of the channel, rather than a change in the C-type inactivation conformational change. We have found that a decrease in the potassium affinity of the internal ion binding site in the pore results in lowered (electrostatic) interactions among ions in the pore and as a result prolongs the time an ion remains bound at the external C-type inactivation site. We also present evidence that the C-type inactivation constriction is quite local and does not involve a general collapse of the selectivity filter. Our data indicate that in A463C potassium can bind within the selectivity filter without interfering with the process of C-type inactivation.

Show MeSH
Related in: MedlinePlus