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Hydrophobic interactions between the voltage sensor and pore mediate inactivation in Kv11.1 channels.

Perry MD, Wong S, Ng CA, Vandenberg JI - J. Gen. Physiol. (2013)

Bottom Line: Crucially in Kv11.1 channels, inactivation gating occurs much more rapidly, and over a distinct range of voltages, compared with activation gating.Combining REFER analysis with double mutant cycle analysis, we provide evidence for a hydrophobic interaction between residues on the S4 and S5 helices.Based on a Kv11.1 channel homology model, we propose that this hydrophobic interaction forms the basis of an intersubunit coupling between the voltage sensor and pore domain that is an important mediator of inactivation gating.

View Article: PubMed Central - HTML - PubMed

Affiliation: Molecular Cardiology and Biophysics Division, Victor Chang Cardiac Research Institute, Darlinghurst, NSW 2010, Australia.

ABSTRACT
Kv11.1 channels are critical for the maintenance of a normal heart rhythm. The flow of potassium ions through these channels is controlled by two voltage-regulated gates, termed "activation" and "inactivation," located at opposite ends of the pore. Crucially in Kv11.1 channels, inactivation gating occurs much more rapidly, and over a distinct range of voltages, compared with activation gating. Although it is clear that the fourth transmembrane segments (S4), within each subunit of the tetrameric channel, are important for controlling the opening and closing of the activation gate, their role during inactivation gating is much less clear. Here, we use rate equilibrium free energy relationship (REFER) analysis to probe the contribution of the S4 "voltage-sensor" helix during inactivation of Kv11.1 channels. Contrary to the important role that charged residues play during activation gating, it is the hydrophobic residues (Leu529, Leu530, Leu532, and Val535) that are the key molecular determinants of inactivation gating. Within the context of an interconnected multi-domain model of Kv11.1 inactivation gating, our REFER analysis indicates that the S4 helix and the S4-S5 linker undergo a conformational rearrangement shortly after that of the S5 helix and S5P linker, but before the S6 helix. Combining REFER analysis with double mutant cycle analysis, we provide evidence for a hydrophobic interaction between residues on the S4 and S5 helices. Based on a Kv11.1 channel homology model, we propose that this hydrophobic interaction forms the basis of an intersubunit coupling between the voltage sensor and pore domain that is an important mediator of inactivation gating.

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Perturbations to inactivation gating and activation gating show poor correlation for S4 residue mutations. Plot comparing perturbations to ΔG0, versus WT, of inactivation gating (ΔΔG0Inact, represented by Δlog(Keq,0)) compared with perturbations to ΔG0, versus WT, of activation gating (ΔΔG0Activ; see Materials and methods) for each S4 residue mutation. Solid black line represents linear regression analysis constrained to go through WT, whereas dashed gray line is an unconstrained fit (R2 = 0.38).
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fig6: Perturbations to inactivation gating and activation gating show poor correlation for S4 residue mutations. Plot comparing perturbations to ΔG0, versus WT, of inactivation gating (ΔΔG0Inact, represented by Δlog(Keq,0)) compared with perturbations to ΔG0, versus WT, of activation gating (ΔΔG0Activ; see Materials and methods) for each S4 residue mutation. Solid black line represents linear regression analysis constrained to go through WT, whereas dashed gray line is an unconstrained fit (R2 = 0.38).

Mentions: Inactivation of Kv11.1 channels is thought to be intrinsically voltage dependent; i.e., the voltage dependence of inactivation is not directly linked to the voltage dependence of activation (Vandenberg et al., 2012). There are two ways that a mutation can affect the voltage-dependent inactivation of Kv11.1 channels. The first is by altering the chemical free energy change associated with inactivation, i.e., a shift in the equilibrium set point of the voltage dependence (compared with WT), as reflected by a change in log(Keq,0). The vast majority of point mutations in the S4 helix (66 out of 70; see Fig. 6 and Table S1), and throughout the channel protein (Wang et al., 2011), caused a negative shift in log(Keq,0), reflecting a depolarizing shift in the voltage dependence of inactivation (i.e., stabilization of the open state). In contrast, the same set of S4 mutations altered the chemical free energy, and hence the voltage dependence, of activation gating in either direction to an approximately equal degree (Fig. 6). Thus, the S4 mutation-induced perturbations to the voltage dependence of inactivation show only a weak correlation (R2 = 0.38) with the perturbations to the voltage dependence of activation (Fig. 6), consistent with the notion that the activation and inactivation gating processes are not directly linked.


Hydrophobic interactions between the voltage sensor and pore mediate inactivation in Kv11.1 channels.

Perry MD, Wong S, Ng CA, Vandenberg JI - J. Gen. Physiol. (2013)

Perturbations to inactivation gating and activation gating show poor correlation for S4 residue mutations. Plot comparing perturbations to ΔG0, versus WT, of inactivation gating (ΔΔG0Inact, represented by Δlog(Keq,0)) compared with perturbations to ΔG0, versus WT, of activation gating (ΔΔG0Activ; see Materials and methods) for each S4 residue mutation. Solid black line represents linear regression analysis constrained to go through WT, whereas dashed gray line is an unconstrained fit (R2 = 0.38).
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC3753607&req=5

fig6: Perturbations to inactivation gating and activation gating show poor correlation for S4 residue mutations. Plot comparing perturbations to ΔG0, versus WT, of inactivation gating (ΔΔG0Inact, represented by Δlog(Keq,0)) compared with perturbations to ΔG0, versus WT, of activation gating (ΔΔG0Activ; see Materials and methods) for each S4 residue mutation. Solid black line represents linear regression analysis constrained to go through WT, whereas dashed gray line is an unconstrained fit (R2 = 0.38).
Mentions: Inactivation of Kv11.1 channels is thought to be intrinsically voltage dependent; i.e., the voltage dependence of inactivation is not directly linked to the voltage dependence of activation (Vandenberg et al., 2012). There are two ways that a mutation can affect the voltage-dependent inactivation of Kv11.1 channels. The first is by altering the chemical free energy change associated with inactivation, i.e., a shift in the equilibrium set point of the voltage dependence (compared with WT), as reflected by a change in log(Keq,0). The vast majority of point mutations in the S4 helix (66 out of 70; see Fig. 6 and Table S1), and throughout the channel protein (Wang et al., 2011), caused a negative shift in log(Keq,0), reflecting a depolarizing shift in the voltage dependence of inactivation (i.e., stabilization of the open state). In contrast, the same set of S4 mutations altered the chemical free energy, and hence the voltage dependence, of activation gating in either direction to an approximately equal degree (Fig. 6). Thus, the S4 mutation-induced perturbations to the voltage dependence of inactivation show only a weak correlation (R2 = 0.38) with the perturbations to the voltage dependence of activation (Fig. 6), consistent with the notion that the activation and inactivation gating processes are not directly linked.

Bottom Line: Crucially in Kv11.1 channels, inactivation gating occurs much more rapidly, and over a distinct range of voltages, compared with activation gating.Combining REFER analysis with double mutant cycle analysis, we provide evidence for a hydrophobic interaction between residues on the S4 and S5 helices.Based on a Kv11.1 channel homology model, we propose that this hydrophobic interaction forms the basis of an intersubunit coupling between the voltage sensor and pore domain that is an important mediator of inactivation gating.

View Article: PubMed Central - HTML - PubMed

Affiliation: Molecular Cardiology and Biophysics Division, Victor Chang Cardiac Research Institute, Darlinghurst, NSW 2010, Australia.

ABSTRACT
Kv11.1 channels are critical for the maintenance of a normal heart rhythm. The flow of potassium ions through these channels is controlled by two voltage-regulated gates, termed "activation" and "inactivation," located at opposite ends of the pore. Crucially in Kv11.1 channels, inactivation gating occurs much more rapidly, and over a distinct range of voltages, compared with activation gating. Although it is clear that the fourth transmembrane segments (S4), within each subunit of the tetrameric channel, are important for controlling the opening and closing of the activation gate, their role during inactivation gating is much less clear. Here, we use rate equilibrium free energy relationship (REFER) analysis to probe the contribution of the S4 "voltage-sensor" helix during inactivation of Kv11.1 channels. Contrary to the important role that charged residues play during activation gating, it is the hydrophobic residues (Leu529, Leu530, Leu532, and Val535) that are the key molecular determinants of inactivation gating. Within the context of an interconnected multi-domain model of Kv11.1 inactivation gating, our REFER analysis indicates that the S4 helix and the S4-S5 linker undergo a conformational rearrangement shortly after that of the S5 helix and S5P linker, but before the S6 helix. Combining REFER analysis with double mutant cycle analysis, we provide evidence for a hydrophobic interaction between residues on the S4 and S5 helices. Based on a Kv11.1 channel homology model, we propose that this hydrophobic interaction forms the basis of an intersubunit coupling between the voltage sensor and pore domain that is an important mediator of inactivation gating.

Show MeSH
Related in: MedlinePlus