Limits...
Direct evidence of conformational changes associated with voltage gating in a voltage sensor protein by time-resolved X-ray/neutron interferometry.

Tronin AY, Nordgren CE, Strzalka JW, Kuzmenko I, Worcester DL, Lauter V, Freites JA, Tobias DJ, Blasie JK - Langmuir (2014)

Bottom Line: Proteins homologous with the VSD have recently been found to function themselves as voltage-gated proton channels or to impart voltage sensitivity to enzymes.Determining the conformational changes associated with voltage gating in the VSD itself in the absence of a pore domain thereby gains importance.The characteristics of the changes observed were in qualitative agreement with molecular dynamics simulations of a related membrane system, suggesting an initial interpretation of these changes in terms of the VSD's atomic-level 3-D structure.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, University of Pennsylvania , Philadelphia, Pennsylvania 19104, United States.

ABSTRACT
The voltage sensor domain (VSD) of voltage-gated cation (e.g., Na(+), K(+)) channels central to neurological signal transmission can function as a distinct module. When linked to an otherwise voltage-insensitive, ion-selective membrane pore, the VSD imparts voltage sensitivity to the channel. Proteins homologous with the VSD have recently been found to function themselves as voltage-gated proton channels or to impart voltage sensitivity to enzymes. Determining the conformational changes associated with voltage gating in the VSD itself in the absence of a pore domain thereby gains importance. We report the direct measurement of changes in the scattering-length density (SLD) profile of the VSD protein, vectorially oriented within a reconstituted phospholipid bilayer membrane, as a function of the transmembrane electric potential by time-resolved X-ray and neutron interferometry. The changes in the experimental SLD profiles for both polarizing and depolarizing potentials with respect to zero potential were found to extend over the entire length of the isolated VSD's profile structure. The characteristics of the changes observed were in qualitative agreement with molecular dynamics simulations of a related membrane system, suggesting an initial interpretation of these changes in terms of the VSD's atomic-level 3-D structure.

Show MeSH

Related in: MedlinePlus

Left side: typical modulus /F(Qz)/ data (top), calculatedfrom the specularX-ray reflectivity R(Qz)/RF(Qz), where /F(Qz)/2 = R(Qz)/RF(Qz), forthe VSD:POPC membrane tethered to the surface of a SiGeSi multilayersubstrate at a transmembrane potential of 0 mV for the first two cyclesof the series of potentials applied. The difference modulus data Δ/F(Qz)/ for the second cycle are shown in the panels belowfor the pairs of potentials indicated. Right side: typical modulus/F(Qz)/ data (top) for the OTS:POPC hybrid bilayer tethered to the surfaceof a SiGeSi multilayer substrate at a transmembrane potential of 0mV for the first two cycles of the series of potentials applied. The difference modulus data Δ/F(Qz)/ for the first cycle areshown in the panels below for the pairs of potentials indicated. Thestandard errors in the data are indicated for each case.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC4007984&req=5

fig2: Left side: typical modulus /F(Qz)/ data (top), calculatedfrom the specularX-ray reflectivity R(Qz)/RF(Qz), where /F(Qz)/2 = R(Qz)/RF(Qz), forthe VSD:POPC membrane tethered to the surface of a SiGeSi multilayersubstrate at a transmembrane potential of 0 mV for the first two cyclesof the series of potentials applied. The difference modulus data Δ/F(Qz)/ for the second cycle are shown in the panels belowfor the pairs of potentials indicated. Right side: typical modulus/F(Qz)/ data (top) for the OTS:POPC hybrid bilayer tethered to the surfaceof a SiGeSi multilayer substrate at a transmembrane potential of 0mV for the first two cycles of the series of potentials applied. The difference modulus data Δ/F(Qz)/ for the first cycle areshown in the panels below for the pairs of potentials indicated. Thestandard errors in the data are indicated for each case.

Mentions: Typical modulusdata /F(Qz)/ from a VSD:POPC membrane tethered to the surface of a SiGeSimultilayer substrate, prepared via “directed assembly (DA)”,17 for the first two cycles of variation of thetransmembrane potential are shown in Figure 2. The potential dependence of the modulus data was used for subsequentanalysis. Since the difference in the modulus databetween p100 mV or m100 mV (“p”/“m” denote+/–, respectively) and 0 mV were similar using the data foreither the first or last 0 mV potential in each cycle, the data forthe two 0 mV values were averaged. The difference modulus data for {p100mV-0mVave}, {m100mV-0mVave}, and {p100mV-m100mV}are also shown in Figure 3. Such differencemodulus data were similar for the first two cycles of the transmembranepotential and are significant because the difference data exceed thestandard errors and they depend on the particular pair of potentialsutilized. Comparable difference modulus data werealso obtained from a VSD:POPC membrane, prepared via “selfassembly (SA)”,17 for the firsttwo cycles of variation of the transmembrane potential, thereby providingadditional support for the reproducibility of these data specimen-to-specimen.By the third and fourth cycles, some evolution of the modulus databecame evident, possibly arising from radiation damage; the analysesof these data will therefore not be presented here. A hybrid bilayermembrane, composed of a chemisorbed layer of OTS with an overlayerof POPC lacking the VSD protein, was employed in this work primarilyonly as a control. Typical modulus data for F(Qz) fromthe OTS:POPC hybrid bilayer for the first two cycles of variationof the transmembrane potential are shown in Figure 2. The difference modulus data for {p100mV-0mVave}and {m100mV-0mVave} are also shown in Figure 2. These difference modulus data were also similar for the first twocycles of the transmembrane potential and are significant becausethe difference data exceed the standard errors and they depend onthe particular pair of potentials utilized. The difference modulusdata for the OTS:POPC hybrid bilayer differ dramatically from thosefor the VSD:POPC membrane, thereby providing an important controlin momentum transfer space.


Direct evidence of conformational changes associated with voltage gating in a voltage sensor protein by time-resolved X-ray/neutron interferometry.

Tronin AY, Nordgren CE, Strzalka JW, Kuzmenko I, Worcester DL, Lauter V, Freites JA, Tobias DJ, Blasie JK - Langmuir (2014)

Left side: typical modulus /F(Qz)/ data (top), calculatedfrom the specularX-ray reflectivity R(Qz)/RF(Qz), where /F(Qz)/2 = R(Qz)/RF(Qz), forthe VSD:POPC membrane tethered to the surface of a SiGeSi multilayersubstrate at a transmembrane potential of 0 mV for the first two cyclesof the series of potentials applied. The difference modulus data Δ/F(Qz)/ for the second cycle are shown in the panels belowfor the pairs of potentials indicated. Right side: typical modulus/F(Qz)/ data (top) for the OTS:POPC hybrid bilayer tethered to the surfaceof a SiGeSi multilayer substrate at a transmembrane potential of 0mV for the first two cycles of the series of potentials applied. The difference modulus data Δ/F(Qz)/ for the first cycle areshown in the panels below for the pairs of potentials indicated. Thestandard errors in the data are indicated for each case.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4007984&req=5

fig2: Left side: typical modulus /F(Qz)/ data (top), calculatedfrom the specularX-ray reflectivity R(Qz)/RF(Qz), where /F(Qz)/2 = R(Qz)/RF(Qz), forthe VSD:POPC membrane tethered to the surface of a SiGeSi multilayersubstrate at a transmembrane potential of 0 mV for the first two cyclesof the series of potentials applied. The difference modulus data Δ/F(Qz)/ for the second cycle are shown in the panels belowfor the pairs of potentials indicated. Right side: typical modulus/F(Qz)/ data (top) for the OTS:POPC hybrid bilayer tethered to the surfaceof a SiGeSi multilayer substrate at a transmembrane potential of 0mV for the first two cycles of the series of potentials applied. The difference modulus data Δ/F(Qz)/ for the first cycle areshown in the panels below for the pairs of potentials indicated. Thestandard errors in the data are indicated for each case.
Mentions: Typical modulusdata /F(Qz)/ from a VSD:POPC membrane tethered to the surface of a SiGeSimultilayer substrate, prepared via “directed assembly (DA)”,17 for the first two cycles of variation of thetransmembrane potential are shown in Figure 2. The potential dependence of the modulus data was used for subsequentanalysis. Since the difference in the modulus databetween p100 mV or m100 mV (“p”/“m” denote+/–, respectively) and 0 mV were similar using the data foreither the first or last 0 mV potential in each cycle, the data forthe two 0 mV values were averaged. The difference modulus data for {p100mV-0mVave}, {m100mV-0mVave}, and {p100mV-m100mV}are also shown in Figure 3. Such differencemodulus data were similar for the first two cycles of the transmembranepotential and are significant because the difference data exceed thestandard errors and they depend on the particular pair of potentialsutilized. Comparable difference modulus data werealso obtained from a VSD:POPC membrane, prepared via “selfassembly (SA)”,17 for the firsttwo cycles of variation of the transmembrane potential, thereby providingadditional support for the reproducibility of these data specimen-to-specimen.By the third and fourth cycles, some evolution of the modulus databecame evident, possibly arising from radiation damage; the analysesof these data will therefore not be presented here. A hybrid bilayermembrane, composed of a chemisorbed layer of OTS with an overlayerof POPC lacking the VSD protein, was employed in this work primarilyonly as a control. Typical modulus data for F(Qz) fromthe OTS:POPC hybrid bilayer for the first two cycles of variationof the transmembrane potential are shown in Figure 2. The difference modulus data for {p100mV-0mVave}and {m100mV-0mVave} are also shown in Figure 2. These difference modulus data were also similar for the first twocycles of the transmembrane potential and are significant becausethe difference data exceed the standard errors and they depend onthe particular pair of potentials utilized. The difference modulusdata for the OTS:POPC hybrid bilayer differ dramatically from thosefor the VSD:POPC membrane, thereby providing an important controlin momentum transfer space.

Bottom Line: Proteins homologous with the VSD have recently been found to function themselves as voltage-gated proton channels or to impart voltage sensitivity to enzymes.Determining the conformational changes associated with voltage gating in the VSD itself in the absence of a pore domain thereby gains importance.The characteristics of the changes observed were in qualitative agreement with molecular dynamics simulations of a related membrane system, suggesting an initial interpretation of these changes in terms of the VSD's atomic-level 3-D structure.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, University of Pennsylvania , Philadelphia, Pennsylvania 19104, United States.

ABSTRACT
The voltage sensor domain (VSD) of voltage-gated cation (e.g., Na(+), K(+)) channels central to neurological signal transmission can function as a distinct module. When linked to an otherwise voltage-insensitive, ion-selective membrane pore, the VSD imparts voltage sensitivity to the channel. Proteins homologous with the VSD have recently been found to function themselves as voltage-gated proton channels or to impart voltage sensitivity to enzymes. Determining the conformational changes associated with voltage gating in the VSD itself in the absence of a pore domain thereby gains importance. We report the direct measurement of changes in the scattering-length density (SLD) profile of the VSD protein, vectorially oriented within a reconstituted phospholipid bilayer membrane, as a function of the transmembrane electric potential by time-resolved X-ray and neutron interferometry. The changes in the experimental SLD profiles for both polarizing and depolarizing potentials with respect to zero potential were found to extend over the entire length of the isolated VSD's profile structure. The characteristics of the changes observed were in qualitative agreement with molecular dynamics simulations of a related membrane system, suggesting an initial interpretation of these changes in terms of the VSD's atomic-level 3-D structure.

Show MeSH
Related in: MedlinePlus