Atomic scale movement of the voltage-sensing region in a potassium channel measured via spectroscopy

Abstract

Voltage-gated ion channels are transmembrane proteins that are essential for nerve impulses and regulate ion flow across cell membranes in response to changes in membrane potential. They are made up of four homologous domains or subunits, each of which contains six transmembrane segments. Studies of potassium channels have shown that the second (S2) and fourth (S4) segments contain several charged residues, which sense changes in voltage and form part of the voltage sensor. Although these regions clearly undergo conformational changes in response to voltage, little is known about the nature of these changes because voltage-dependent distance changes have not been measured. Here we use lanthanide-based resonance energy transfer to measure distances between Shaker potassium channel subunits at specific residues. Voltage-dependent distance changes of up to 3.2 A were measured at several sites near the S4 segment. These movements directly correlated with electrical measurements of the voltage sensor, establishing the link between physical changes and electrical charge movement. Measured distance changes suggest that the region associated with the S4 segment undergoes a rotation and possible tilt, rather than a large transmembrane movement, in response to voltage. These results demonstrate the first in situ measurement of atomic scale movement in a trans-membrane protein.

S4 Charges Move Close to Residues in the Pore Domain during Activation in a K Channel

“…This confirms earlier suggestions of the location of S4 relative to the pore (Gandhi et al 2000; Elinder et al 2001). The large motion of R362 suggested by our results is compatible with both a helical screw motion (Catterall 1986; Guy and Seetharamulu 1986; Glauner et al 1999; Keynes and Elinder 1999; Gandhi et al 2000) and a pure twisting motion (Papazian and Bezanilla 1997; Cha et al 1999; Glauner et al 1999) of S4. Our estimated location and motion is also compatible with the results of other studies (Li-Smerin et al 2000; Loots and Isacoff 2000).…”

“…Cysteine accessibility studies suggest that the positive charges of S4 are either buried in the membrane or in the cytosol when the membrane is held at a hyperpolarized potential, and, in response to a depolarizing voltage pulse, the S4 charges move outward and exposes its three most external charges into the extracellular solution (Larsson et al 1996; Yang et al 1996; Yusaf et al 1996; Baker et al 1998; for review see Keynes and Elinder 1999). S4 is suggested to undergo a large-scale movement during activation of the channels, either a 180° rotation (Papazian and Bezanilla 1997; Cha et al 1999; Glauner et al 1999) or a helical screw motion with both a 180° rotation and a translational motion of 13.5 Å (Catterall 1986; Guy and Seetharamulu 1986; Glauner et al 1999; Keynes and Elinder 1999; Gandhi et al 2000). In a recent study on the Shaker K channel, we suggested that S4, in the activated state, was located just outside S5 with the most external positive charge of S4 (R362) closest to the pore (Elinder et al 2001).…”

PolydA and polyrA self‐structured by a europium and amino acid complex

“…Circular dichroism spectra (CD) and CD melting experiments were measured on a JASCO J-810 spectropolarimeter equipped with a temperature-controlled water-bath. 1 H NMR spectra were carried out on a Bruker Avance 600 MHz NMR Spectrometer in D 2 O at 20°C.…”

“…Due to a unique 4f n electronic configuration, lanthanide materials have been widely used as probes in luminescent resonance energy transfer (LRET) for bioassays and as reagents for diagnosis in magnetic resonance imaging (MRI) [1][2][3][4][5]. As chemical nucleases, lanthanide complexes have also shown a high efficiency to hydrolyze DNA and RNA without redox chemistry [6,7] There is great interest in the design and synthesis of small molecules which selectively target specific genes to inhibit biological functions in which particular DNA structures participate [8][9][10][11][12][13][14][15].…”

Functional diversity of potassium channel voltage-sensing domains

“…49,50 Later measurements using spectroscopic techniques, particularly the LRET and another variation of FRET involving transfer between dyes and hydrophobic quenchers, suggested that the magnitude of movement was at most a couple of Angstroms (A ). 51,52 In contrast, a proposal based on the KvAP structure and state-dependent avidin/biotin accessibility experiments, 53 suggested that the S3-S4, therein named the paddle, might traverse most of the membrane thickness in a rigid body motion. Although there is not a consensus, most models of voltage-dependent gating seem to suggest that the S4 segment translates several angstroms and undergoes a~120 degree counter clockwise rotation.…”