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Structural changes in single membranes in response to an applied transmembrane electric potential revealed by time-resolved neutron/X-ray interferometry.

ABSTRACT: The profile structure of a hybrid lipid bilayer, tethered to the surface of an inorganic substrate and fully hydrated with a bulk aqueous medium in an electrochemical cell, was investigated as a function of the applied transbilayer electric potential via time-resolved neutron reflectivity, enhanced by interferometry. Significant, and fully reversible structural changes were observed in the distal half (with respect to the substrate surface) of the hybrid bilayer comprised of a zwitterionic phospholipid in response to a +100mV potential with respect to 0mV. These arise presumably due to reorientation of the electric dipole present in the polar headgroup of the phospholipid and its resulting effect on the thickness of the phospholipid's hydrocarbon chain layer within the hybrid bilayer's profile structure. The profile structure of the voltage-sensor domain from a voltage-gated ion channel protein within a phospholipid bilayer membrane, tethered to the surface of an inorganic substrate and fully hydrated with a bulk aqueous medium in an electrochemical cell, was also investigated as a function of the applied transmembrane electric potential via time-resolved X-ray reflectivity, enhanced by interferometry. Significant, fully-reversible, and different structural changes in the protein were detected in response to ±100mV potentials with respect to 0mV. The approach employed is that typical of transient spectroscopy, shown here to be applicable to both neutron and X-ray reflectivity of thin films.

SUBMITTER: Tronin A

PROVIDER: S-EPMC3821765 | biostudies-literature | 2013 Aug

REPOSITORIES: biostudies-literature

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Structural changes in single membranes in response to an applied transmembrane electric potential revealed by time-resolved neutron/X-ray interferometry.

Tronin A A Chen C-H CH Gupta S S Worcester D D Lauter V V Strzalka J J Kuzmenko I I Blasie J K JK

Chemical physics 20130801

The profile structure of a hybrid lipid bilayer, tethered to the surface of an inorganic substrate and fully hydrated with a bulk aqueous medium in an electrochemical cell, was investigated as a function of the applied transbilayer electric potential via time-resolved neutron reflectivity, enhanced by interferometry. Significant, and fully reversible structural changes were observed in the distal half (with respect to the substrate surface) of the hybrid bilayer comprised of a zwitterionic phosp ...[more]

PMID: 24222930

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Project description:Available experimental techniques cannot determine high-resolution three-dimensional structures of membrane proteins under a transmembrane voltage. Hence, the mechanism by which voltage-gated cation channels couple conformational changes within the four voltage sensor domains, in response to either depolarizing or polarizing transmembrane voltages, to opening or closing of the pore domain's ion channel remains unresolved. Single-membrane specimens, composed of a phospholipid bilayer containing a vectorially oriented voltage-gated K+ channel protein at high in-plane density tethered to the surface of an inorganic multilayer substrate, were developed to allow the application of transmembrane voltages in an electrochemical cell. Time-resolved neutron reflectivity experiments, enhanced by interferometry enabled by the multilayer substrate, were employed to provide directly the low-resolution profile structures of the membrane containing the vectorially oriented voltage-gated K+ channel for the activated, open and deactivated, closed states of the channel under depolarizing and hyperpolarizing transmembrane voltages applied cyclically. The profile structures of these single membranes were dominated by the voltage-gated K+ channel protein because of the high in-plane density. Importantly, the use of neutrons allowed the determination of the voltage-dependent changes in both the profile structure of the membrane and the distribution of water within the profile structure. These two key experimental results were then compared to those predicted by three computational modeling approaches for the activated, open and deactivated, closed states of three different voltage-gated K+ channels in hydrated phospholipid bilayer membrane environments. Of the three modeling approaches investigated, only one state-of-the-art molecular dynamics simulation that directly predicted the response of a voltage-gated K+ channel within a phospholipid bilayer membrane to applied transmembrane voltages by utilizing very long trajectories was found to be in agreement with the two key experimental results provided by the time-resolved neutron interferometry experiments.

Project description:One of the most important questions in biological science is how a protein functions. When a protein performs its function, it undergoes regulated structural transitions. In this regard, to better understand the underlying principle of a protein function, it is desirable to monitor the dynamic evolution of the protein structure in real time. To probe fast and subtle motions of a protein in physiological conditions demands an experimental tool that is not only equipped with superb spatiotemporal resolution but also applicable to samples in solution phase. Time-resolved X-ray solution scattering (TRXSS), discussed in this Account, fits all of those requirements needed for probing the movements of proteins in aqueous solution. The technique utilizes a pump-probe scheme employing an optical pump pulse to initiate photoreactions of proteins and an X-ray probe pulse to monitor ensuing structural changes. The technical advances in ultrafast lasers and X-ray sources allow us to achieve superb temporal resolution down to femtoseconds. Because X-rays scatter off all atomic pairs in a protein, an X-ray scattering pattern provides information on the global structure of the protein with subangstrom spatial resolution. Importantly, TRXSS is readily applicable to aqueous solution samples of proteins with the aid of theoretical models and therefore is well suited for investigating structural dynamics of protein transitions in physiological conditions. In this Account, we demonstrate that TRXSS can be used to probe real-time structural dynamics of proteins in solution ranging from subtle helix movement to global conformational change. Specifically, we discuss the photoreactions of photoactive yellow protein (PYP) and homodimeric hemoglobin (HbI). For PYP, we revealed the kinetics of structural transitions among four transient intermediates comprising a photocycle and, by applying structural analysis based on ab initio shape reconstruction, showed that the signaling of PYP involves the protrusion of the N-terminus with significant increase of the overall protein size. For HbI, we elucidated the dynamics of complex allosteric transitions among transient intermediates. In particular, by applying structural refinement analysis based on rigid-body modeling, we found that the allosteric transition of HbI accompanies the rotation of quaternary structure and the contraction between two heme domains. By making use of the experimental and analysis methods presented in this Account, we envision that the TRXSS can be used to probe the structural dynamics of various proteins, allowing us to decipher the working mechanisms of their functions. Furthermore, when combined with femtosecond X-ray pulses generated from X-ray free electron lasers, TRXSS will gain access to ultrafast protein dynamics on sub-picosecond time scales.

Dataset Information

Structural changes in single membranes in response to an applied transmembrane electric potential revealed by time-resolved neutron/X-ray interferometry.

Publications

Structural changes in single membranes in response to an applied transmembrane electric potential revealed by time-resolved neutron/X-ray interferometry.

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