Amphiphilic blockers punch through a mutant CLC-0 pore.

Intracellularly applied amphiphilic molecules, such as p-chlorophenoxy acetate (CPA) and octanoate, block various pore-open mutants of CLC-0. The voltage-dependent block of a particular pore-open mutant, E166G, was found to be multiphasic. In symmetrical 140 mM Cl(-), the apparent affinity of the blocker in this mutant increased with a negative membrane potential but, paradoxically, decreased when the negative membrane potential was greater than -80 mV, a phenomenon similar to the blocker "punch-through" shown in many blocker studies of cation channels. To provide further evidence of the punch-through of CPA and octanoate, we studied the dissociation rate of the blocker from the pore by measuring the time constant of relief from the block under various voltage and ionic conditions. Consistent with the voltage dependence of the effect on the steady-state current, the rate of CPA dissociation from the E166G pore reached a minimum at -80 mV in symmetrical 140 mM Cl(-), and the direction of current recovery suggested that the bound CPA in the pore can dissociate into both intracellular and extracellular solutions. Moreover, the CPA dissociation depends upon the Cl(-) reversal potential with a minimal dissociation rate at a voltage 80 mV more negative than the Cl(-) reversal potential. That the shift of the CPA-dissociation rate follows the Cl(-) gradient across the membrane argues that these blockers can indeed punch through the channel pore. Furthermore, a minimal CPA-dissociation rate at a voltage 80 mV more negative than the Cl(-) reversal potential suggests that the outward blocker movement through the CLC-0 pore is more difficult than the inward movement

Modulation of the slow/common gating of CLC channels by intracellular cadmium.

Members of the CLC family of Cl(-) channels and transporters are homodimeric integral membrane proteins. Two gating mechanisms control the opening and closing of Cl(-) channels in this family: fast gating, which regulates opening and closing of the individual pores in each subunit, and slow (or common) gating, which simultaneously controls gating of both subunits. Here, we found that intracellularly applied Cd(2+) reduces the current of CLC-0 because of its inhibition on the slow gating. We identified CLC-0 residues C229 and H231, located at the intracellular end of the transmembrane domain near the dimer interface, as the Cd(2+)-coordinating residues. The inhibition of the current of CLC-0 by Cd(2+) was greatly enhanced by mutation of I225W and V490W at the dimer interface. Biochemical experiments revealed that formation of a disulfide bond within this Cd(2+)-binding site is also affected by mutation of I225W and V490W, indicating that these two mutations alter the structure of the Cd(2+)-binding site. Kinetic studies showed that Cd(2+) inhibition appears to be state dependent, suggesting that structural rearrangements may occur in the CLC dimer interface during Cd(2+) modulation. Mutations of I290 and I556 of CLC-1, which correspond to I225 and V490 of CLC-0, respectively, have been shown previously to cause malfunction of CLC-1 Cl(-) channel by altering the common gating. Our experimental results suggest that mutations of the corresponding residues in CLC-0 change the subunit interaction and alter the slow gating of CLC-0. The effect of these mutations on modulations of slow gating of CLC channels by intracellular Cd(2+) likely depends on their alteration of subunit interactions

Mixing and combining with AOA and TOA for the enhanced accuracy of mobile location

[[abstract]]Position location has become a hot issue over the past few years in wireless communication. Providing the accurate location information of the mobile station (MS) is necessitated by the Emergent 911 call in United States. The angle of arrival (AOA), time of arrival (TOA), and time difference of arrival (TDOA) techniques have been proposed for providing location services in wireless networks. We present a method for the enhanced accuracy of mobile location. This method is mixing and combining with AOA and TOA in wireless networks and picks out mobile location with large deviation to enhance the accuracy of location estimation. Numerical results demonstrate that the proposed location scheme gives much higher location accuracy than the method that only used TOA and AOA location technique.[[notice]]需補地點及國別[[conferencetype]]國際[[conferencedate]]20030422~2003042

Side-chain Charge Effects and Conductance Determinants in the Pore of ClC-0 Chloride Channels

The charge on the side chain of the internal pore residue lysine 519 (K519) of the Torpedo ClC-0 chloride (Cl−) channel affects channel conductance. Experiments that replace wild-type (WT) lysine with neutral or negatively charged residues or that modify the K519C mutant with various methane thiosulfonate (MTS) reagents show that the conductance of the channel decreases when the charge at position 519 is made more negative. This charge effect on the channel conductance diminishes in the presence of a high intracellular Cl− concentration ([Cl−]i). However, the application of high concentrations of nonpermeant ions, such as glutamate or sulfate (SO42−), does not change the conductance, suggesting that the electrostatic effects created by the charge at position 519 are unlikely due to a surface charge mechanism. Another pore residue, glutamate 127 (E127), plays an even more critical role in controlling channel conductance. This negatively charged residue, based on the structures of the homologous bacterial ClC channels, lies 4–5 Å from K519. Altering the charge of this residue can influence the apparent Cl− affinity as well as the saturated pore conductance in the conductance-Cl− activity curve. Amino acid residues at the selectivity filter also control the pore conductance but mutating these residues mainly affects the maximal pore conductance. These results suggest at least two different conductance determinants in the pore of ClC-0, consistent with the most recent crystal structure of the bacterial ClC channel solved to 2.5 Å, in which multiple Cl−-binding sites were identified in the pore. Thus, we suggest that the occupancy of the internal Cl−-binding site is directly controlled by the charged residues located at the inner pore mouth. On the other hand, the Cl−-binding site at the selectivity filter controls the exit rate of Cl− and therefore determines the maximal channel conductance

Different Fast-Gate Regulation by External Cl− and H+ of the Muscle-Type Clc Chloride Channels

The fast gate of the muscle-type ClC channels (ClC-0 and ClC-1) opens in response to the change of membrane potential (V). This gating process is intimately associated with the binding of external Cl− to the channel pore in a way that the occupancy of Cl− on the binding site increases the channel's open probability (Po). External H+ also enhances the fast-gate opening in these channels, prompting a hypothesis that protonation of the binding site may increase the Cl− binding affinity, and this is possibly the underlying mechanism for the H+ modulation. However, Cl− and H+, modulate the fast-gate Po-V curve in different ways. Varying the external Cl− concentrations ([Cl−]o) shifts the Po-V curve in parallel along the voltage axis, whereas reducing external pH mainly increases the minimal Po of the curve. Furthermore, H+ modulations at saturating and nonsaturating [Cl−]o are similar. Thus, the H+ effect on the fast gating appears not to be a consequence of an increase in the Cl− binding affinity. We previously found that a hyperpolarization-favored opening process is important to determine the fast-gate Po of ClC-0 at very negative voltages. This [Cl−]o-independent mechanism attracted little attention, but it appears to be the opening process that is modulated by external H+

High Chern number quantum anomalous Hall phases in graphene ribbons with Haldane orbital coupling

We investigate possible phase transitions among the different quantum anomalous Hall (QAH) phases in a zigzag graphene ribbon under the influence of the exchange field. The effective tight-binding Hamiltonian for graphene is made up of the hopping term, the Kane-Mele and Rashba spin-orbit couplings as well as the Haldane orbital term. We find that the variation of the exchange field results in bulk gap-closing phenomena and phase transitions occur in the graphene system. If the Haldane orbital coupling is absent, the phase transition between the chiral (anti-chiral) edge state $\nu=+2$ ($\nu=-2$) and the pseudo-quantum spin Hall state ($\nu=0$) takes place. Surprisingly, when the Haldane orbital coupling is taken into account, an intermediate QSH phase with two additional edge modes appears in between phases $\nu=+2$ and $\nu=-2$. This intermediate phase is therefore either the hyper-chiral edge state of high Chern number $\nu=+4$ or anti-hyper-chiral edge state of $\nu=-4$ when the direction of exchange field is reversed. We present the band structures, edge state wave functions and current distributions of the different QAH phases in the system. We also report the critical exchange field values for the QAH phase transitions.Comment: 4 figure