Although the pathway to the external vestibule can be deduced from the wild-type structure, the task of locating the ion-conducting pathway is simplified by overlaying the E148A postulated open structure. ![]() There are deep depressions on each side of the protein, but atoms from two of the central residues in the EcClC structure, Ser-107 and Glu-148, occlude the pore. There is no continuous course which ions can traverse from the intracellular to extracellular sides of the protein, as shown in Fig. To reveal the ion-conducting path, the front half of the protein is removed in the figure. Later we demonstrate that the results we obtain with the model based on the low-resolution x-ray structure do not differ from those obtained with the model based on the high-resolution structure. ), we choose the wild-type EcClC as our template for this study, due to the presence of the key gating residue Glu-148 in its native state. Moreover, the ion-conducting path of the EcClC channel, unlike that of the KcsA channel ( Thus, before the crystal structure coordinates can be used to investigate the permeation of ions through the channel, a completely open state structure needs to be created by using molecular dynamics or other means. It is likely that the conduction path of ions in these channels has become distorted in the process of crystallization or that they represent the channel in a closed state. coli E148A mutant reveals that two residues are still partially occluding the channel, preventing Cl − permeation. ![]() Investigation of the postulated open state configuration of the E. As we follow the EcClC pore from either the extracellular or intracellular opening toward the middle of the pore, it abruptly tapers and vanishes. coli ClC (EcClC) channel structure, residues from the N-termini of the D, F, and N α-helices are constricting the channel and two of these residues are completely blocking the conduction pore. One of the difficulties in utilizing the newly unveiled information is that all of the published crystallographic structures, including the E148A mutant channel in the postulated open state, have atoms occluding the pore and obstructing Cl − permeation. , 2003), as yet there has been no theoretical study that attempts to relate the atomic structure of a ClC channel to the macroscopic properties. Finally, we demonstrate that a ClC-0 homology model created from an alternative sequence alignment fails to replicate any of the experimental observations. We locate the binding sites, as well as pinpointing the rate-limiting steps in conduction, and make testable predictions about how the single channel current across ClC-0 and ClC-1 will vary as the ionic concentrations are increased. ![]() We find that conduction in these pores involves three ions. Employing open-state ClC-0 and ClC-1 channel models, current-voltage curves consistent with experimental measurements are obtained. Then, retaining the same pore shape, the prokaryotic ClC channel is converted to either ClC-0 or ClC-1 by replacing all the nonconserved dipole-containing and charged amino acid residues. Two residues that are occluding the channel are slowly pushed outward with molecular dynamics to create a continuous ion-conducting path with the minimum radius of 2.5 Å. We create an open-state configuration of the prokaryotic ClC Cl − channel using its known crystallographic structure as a basis. The conduction properties of ClC-0 and ClC-1 chloride channels are examined using electrostatic calculations and three-dimensional Brownian dynamics simulations.
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