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alpha-helical structural elements within the voltage-sensing domains of a K(+) channel.

Li-Smerin Y, Hackos DH, Swartz KJ - J. Gen. Physiol. (2000)

Bottom Line: Our results are consistent with at least portions of S1, S2, S3, and S4 adopting alpha-helical secondary structure.The distribution of gating perturbations for S1 and S2 suggest that these two helices interact primarily with two environments.In contrast, the distribution of perturbations for S3 and S4 were more complex, suggesting that the latter two helices make more extensive protein contacts, possibly interfacing directly with the shell of the pore domain.

View Article: PubMed Central - PubMed

Affiliation: Molecular Physiology and Biophysics Unit, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland 20892, USA.

ABSTRACT
Voltage-gated K(+) channels are tetramers with each subunit containing six (S1-S6) putative membrane spanning segments. The fifth through sixth transmembrane segments (S5-S6) from each of four subunits assemble to form a central pore domain. A growing body of evidence suggests that the first four segments (S1-S4) comprise a domain-like voltage-sensing structure. While the topology of this region is reasonably well defined, the secondary and tertiary structures of these transmembrane segments are not. To explore the secondary structure of the voltage-sensing domains, we used alanine-scanning mutagenesis through the region encompassing the first four transmembrane segments in the drk1 voltage-gated K(+) channel. We examined the mutation-induced perturbation in gating free energy for periodicity characteristic of alpha-helices. Our results are consistent with at least portions of S1, S2, S3, and S4 adopting alpha-helical secondary structure. In addition, both the S1-S2 and S3-S4 linkers exhibited substantial helical character. The distribution of gating perturbations for S1 and S2 suggest that these two helices interact primarily with two environments. In contrast, the distribution of perturbations for S3 and S4 were more complex, suggesting that the latter two helices make more extensive protein contacts, possibly interfacing directly with the shell of the pore domain.

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Distribution of free energy perturbations in channel gating. (A) The bar graph plots the absolute value of ΔΔG0 for all mutations studied. Data are mean ± SEM from Table . The solid line superimposed on the /ΔΔG0/ plot is a windowed hydrophobicity index calculated using the Kyte-Doolittle scale (Kyte and Doolittle 1982) with a 17-residue window. (B) Sliding window analysis for mean ΔΔG0 (black line) and hydrophobicity index (gray line). Both use a 17-residue sliding window. Letters and numbers indicate the wild-type residues and their positions, respectively.
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Figure 3: Distribution of free energy perturbations in channel gating. (A) The bar graph plots the absolute value of ΔΔG0 for all mutations studied. Data are mean ± SEM from Table . The solid line superimposed on the /ΔΔG0/ plot is a windowed hydrophobicity index calculated using the Kyte-Doolittle scale (Kyte and Doolittle 1982) with a 17-residue window. (B) Sliding window analysis for mean ΔΔG0 (black line) and hydrophobicity index (gray line). Both use a 17-residue sliding window. Letters and numbers indicate the wild-type residues and their positions, respectively.

Mentions: The absolute values of ΔΔG0 for all the mutant channels are shown in Fig. 3 superimposed with a Kyte-Doolittle hydrophobicity analysis (Kyte and Doolittle 1982) for the region. Residues with large /ΔΔG0/ values tend to cluster into four groups separated by stretches of relatively small /ΔΔG0/ values. The distribution of perturbation energy correlates nicely with the hydrophobicity profile. The energy maxima correspond to the peaks in the hydrophobicity index, while the energy minima correspond to the valleys in the index. Thus, mutations in the linkers between transmembrane segments where the residues tend to be more hydrophilic typically have much smaller effects on gating. The results are quite remarkable and support the notion that four transmembrane segments are present within this region of voltage-gated K+ channels. While the largest perturbations are seen in S4 (up to 6.6 kcal mol−1), there are also sizable changes in ΔΔG0 for the other three transmembrane segments. It is interesting that there is a trend in the perturbation energies across the region studied with the smallest effects occurring in S1 and the largest effects occurring in S4.


alpha-helical structural elements within the voltage-sensing domains of a K(+) channel.

Li-Smerin Y, Hackos DH, Swartz KJ - J. Gen. Physiol. (2000)

Distribution of free energy perturbations in channel gating. (A) The bar graph plots the absolute value of ΔΔG0 for all mutations studied. Data are mean ± SEM from Table . The solid line superimposed on the /ΔΔG0/ plot is a windowed hydrophobicity index calculated using the Kyte-Doolittle scale (Kyte and Doolittle 1982) with a 17-residue window. (B) Sliding window analysis for mean ΔΔG0 (black line) and hydrophobicity index (gray line). Both use a 17-residue sliding window. Letters and numbers indicate the wild-type residues and their positions, respectively.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC1887781&req=5

Figure 3: Distribution of free energy perturbations in channel gating. (A) The bar graph plots the absolute value of ΔΔG0 for all mutations studied. Data are mean ± SEM from Table . The solid line superimposed on the /ΔΔG0/ plot is a windowed hydrophobicity index calculated using the Kyte-Doolittle scale (Kyte and Doolittle 1982) with a 17-residue window. (B) Sliding window analysis for mean ΔΔG0 (black line) and hydrophobicity index (gray line). Both use a 17-residue sliding window. Letters and numbers indicate the wild-type residues and their positions, respectively.
Mentions: The absolute values of ΔΔG0 for all the mutant channels are shown in Fig. 3 superimposed with a Kyte-Doolittle hydrophobicity analysis (Kyte and Doolittle 1982) for the region. Residues with large /ΔΔG0/ values tend to cluster into four groups separated by stretches of relatively small /ΔΔG0/ values. The distribution of perturbation energy correlates nicely with the hydrophobicity profile. The energy maxima correspond to the peaks in the hydrophobicity index, while the energy minima correspond to the valleys in the index. Thus, mutations in the linkers between transmembrane segments where the residues tend to be more hydrophilic typically have much smaller effects on gating. The results are quite remarkable and support the notion that four transmembrane segments are present within this region of voltage-gated K+ channels. While the largest perturbations are seen in S4 (up to 6.6 kcal mol−1), there are also sizable changes in ΔΔG0 for the other three transmembrane segments. It is interesting that there is a trend in the perturbation energies across the region studied with the smallest effects occurring in S1 and the largest effects occurring in S4.

Bottom Line: Our results are consistent with at least portions of S1, S2, S3, and S4 adopting alpha-helical secondary structure.The distribution of gating perturbations for S1 and S2 suggest that these two helices interact primarily with two environments.In contrast, the distribution of perturbations for S3 and S4 were more complex, suggesting that the latter two helices make more extensive protein contacts, possibly interfacing directly with the shell of the pore domain.

View Article: PubMed Central - PubMed

Affiliation: Molecular Physiology and Biophysics Unit, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland 20892, USA.

ABSTRACT
Voltage-gated K(+) channels are tetramers with each subunit containing six (S1-S6) putative membrane spanning segments. The fifth through sixth transmembrane segments (S5-S6) from each of four subunits assemble to form a central pore domain. A growing body of evidence suggests that the first four segments (S1-S4) comprise a domain-like voltage-sensing structure. While the topology of this region is reasonably well defined, the secondary and tertiary structures of these transmembrane segments are not. To explore the secondary structure of the voltage-sensing domains, we used alanine-scanning mutagenesis through the region encompassing the first four transmembrane segments in the drk1 voltage-gated K(+) channel. We examined the mutation-induced perturbation in gating free energy for periodicity characteristic of alpha-helices. Our results are consistent with at least portions of S1, S2, S3, and S4 adopting alpha-helical secondary structure. In addition, both the S1-S2 and S3-S4 linkers exhibited substantial helical character. The distribution of gating perturbations for S1 and S2 suggest that these two helices interact primarily with two environments. In contrast, the distribution of perturbations for S3 and S4 were more complex, suggesting that the latter two helices make more extensive protein contacts, possibly interfacing directly with the shell of the pore domain.

Show MeSH
Related in: MedlinePlus