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Heme regulates allosteric activation of the Slo1 BK channel.

Horrigan FT, Heinemann SH, Hoshi T - J. Gen. Physiol. (2005)

Bottom Line: At saturating levels of divalent cations, heme remained similarly effective with its influence on the G-V simulated by weakening the coupling of both Ca(2+) binding and voltage sensor activation to channel opening.The results thus show that heme dampens the influence of allosteric activators on the activation gate of the Slo1 BK channel.To account for these effects, we consider the possibility that heme binding alters the structure of the RCK gating ring and thereby disrupts both Ca(2+)- and voltage-dependent gating as well as intrinsic stability of the open state.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, School of Medecine, University of Pennsylvania, Philadelphia, PA 19104, USA.

ABSTRACT
Large conductance calcium-dependent (Slo1 BK) channels are allosterically activated by membrane depolarization and divalent cations, and possess a rich modulatory repertoire. Recently, intracellular heme has been identified as a potent regulator of Slo1 BK channels (Tang, X.D., R. Xu, M.F. Reynolds, M.L. Garcia, S.H. Heinemann, and T. Hoshi. 2003. Nature. 425:531-535). Here we investigated the mechanism of the regulatory action of heme on heterologously expressed Slo1 BK channels by separating the influences of voltage and divalent cations. In the absence of divalent cations, heme generally decreased ionic currents by shifting the channel's G-V curve toward more depolarized voltages and by rendering the curve less steep. In contrast, gating currents remained largely unaffected by heme. Simulations suggest that a decrease in the strength of allosteric coupling between the voltage sensor and the activation gate and a concomitant stabilization of the open state account for the essential features of the heme action in the absence of divalent ions. At saturating levels of divalent cations, heme remained similarly effective with its influence on the G-V simulated by weakening the coupling of both Ca(2+) binding and voltage sensor activation to channel opening. The results thus show that heme dampens the influence of allosteric activators on the activation gate of the Slo1 BK channel. To account for these effects, we consider the possibility that heme binding alters the structure of the RCK gating ring and thereby disrupts both Ca(2+)- and voltage-dependent gating as well as intrinsic stability of the open state.

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Open-channel i–V is not markedly altered by heme. (A) Channel openings elicited by ramp depolarization from 0 to 240 mV in the control condition (left) and with 100 nM heme (right). The sweeps designated by 1 and 2 show the main conductance level. The sweep designated by 3 shows the 60% substate and that by 4 illustrates the 40% substate. (B) Composite i-V curves obtained by conditional averaging of many openings as shown in A.
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fig3: Open-channel i–V is not markedly altered by heme. (A) Channel openings elicited by ramp depolarization from 0 to 240 mV in the control condition (left) and with 100 nM heme (right). The sweeps designated by 1 and 2 show the main conductance level. The sweep designated by 3 shows the 60% substate and that by 4 illustrates the 40% substate. (B) Composite i-V curves obtained by conditional averaging of many openings as shown in A.

Mentions: Despite the dramatic reduction in the macroscopic current described above, the open-channel i–V properties of Slo1 remained largely unaltered. The composite i–V curves for the main conductance state in the control condition and the experimental condition with heme (100 nM; Fig. 3 A, left and right sweeps 1 and 2) were indistinguishable (Fig. 3 B). However, in the presence of heme, smaller conductance levels, corresponding to ∼60 and 40% of the full level (Fig. 3 A, sweeps 3 and 4), were more frequently observed. The increased occurrence of these substates was unequivocal, but the extent of the increase was, however, variable from one patch to the next and difficult to quantify. The rectification properties of these substates were not markedly different from those of the main state. While the occurrence of these substates probably contributed to the macroscopic current inhibition by heme, most of the inhibitory effect of heme was likely mediated by changes in the channel gating as described below.


Heme regulates allosteric activation of the Slo1 BK channel.

Horrigan FT, Heinemann SH, Hoshi T - J. Gen. Physiol. (2005)

Open-channel i–V is not markedly altered by heme. (A) Channel openings elicited by ramp depolarization from 0 to 240 mV in the control condition (left) and with 100 nM heme (right). The sweeps designated by 1 and 2 show the main conductance level. The sweep designated by 3 shows the 60% substate and that by 4 illustrates the 40% substate. (B) Composite i-V curves obtained by conditional averaging of many openings as shown in A.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2266614&req=5

fig3: Open-channel i–V is not markedly altered by heme. (A) Channel openings elicited by ramp depolarization from 0 to 240 mV in the control condition (left) and with 100 nM heme (right). The sweeps designated by 1 and 2 show the main conductance level. The sweep designated by 3 shows the 60% substate and that by 4 illustrates the 40% substate. (B) Composite i-V curves obtained by conditional averaging of many openings as shown in A.
Mentions: Despite the dramatic reduction in the macroscopic current described above, the open-channel i–V properties of Slo1 remained largely unaltered. The composite i–V curves for the main conductance state in the control condition and the experimental condition with heme (100 nM; Fig. 3 A, left and right sweeps 1 and 2) were indistinguishable (Fig. 3 B). However, in the presence of heme, smaller conductance levels, corresponding to ∼60 and 40% of the full level (Fig. 3 A, sweeps 3 and 4), were more frequently observed. The increased occurrence of these substates was unequivocal, but the extent of the increase was, however, variable from one patch to the next and difficult to quantify. The rectification properties of these substates were not markedly different from those of the main state. While the occurrence of these substates probably contributed to the macroscopic current inhibition by heme, most of the inhibitory effect of heme was likely mediated by changes in the channel gating as described below.

Bottom Line: At saturating levels of divalent cations, heme remained similarly effective with its influence on the G-V simulated by weakening the coupling of both Ca(2+) binding and voltage sensor activation to channel opening.The results thus show that heme dampens the influence of allosteric activators on the activation gate of the Slo1 BK channel.To account for these effects, we consider the possibility that heme binding alters the structure of the RCK gating ring and thereby disrupts both Ca(2+)- and voltage-dependent gating as well as intrinsic stability of the open state.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, School of Medecine, University of Pennsylvania, Philadelphia, PA 19104, USA.

ABSTRACT
Large conductance calcium-dependent (Slo1 BK) channels are allosterically activated by membrane depolarization and divalent cations, and possess a rich modulatory repertoire. Recently, intracellular heme has been identified as a potent regulator of Slo1 BK channels (Tang, X.D., R. Xu, M.F. Reynolds, M.L. Garcia, S.H. Heinemann, and T. Hoshi. 2003. Nature. 425:531-535). Here we investigated the mechanism of the regulatory action of heme on heterologously expressed Slo1 BK channels by separating the influences of voltage and divalent cations. In the absence of divalent cations, heme generally decreased ionic currents by shifting the channel's G-V curve toward more depolarized voltages and by rendering the curve less steep. In contrast, gating currents remained largely unaffected by heme. Simulations suggest that a decrease in the strength of allosteric coupling between the voltage sensor and the activation gate and a concomitant stabilization of the open state account for the essential features of the heme action in the absence of divalent ions. At saturating levels of divalent cations, heme remained similarly effective with its influence on the G-V simulated by weakening the coupling of both Ca(2+) binding and voltage sensor activation to channel opening. The results thus show that heme dampens the influence of allosteric activators on the activation gate of the Slo1 BK channel. To account for these effects, we consider the possibility that heme binding alters the structure of the RCK gating ring and thereby disrupts both Ca(2+)- and voltage-dependent gating as well as intrinsic stability of the open state.

Show MeSH
Related in: MedlinePlus