![]() ![]() Furthermore, removing the positive charge at this position reduces and increases, respectively, the accessibility of the negatively and positively charged methane thiosulfonate reagents to the pore. The anion permeability ratios derived from bi-ionic experiments are also significantly altered in that the selectivity of Cl − over other anions is decreased. The mutation of K149 into a neutral residue leucine (K149L) shifts the activation curve and leads to flickery channel openings. On the other hand, mutations of H401 alter the channel conductance but not the gating properties, while mutations of G352 result in very little functional consequence. The experimental results revealed that among the three residues tested, K149 plays the most important role in controlling both the gating and the permeation functions of ClC-0. A cysteine modification approach was also employed to evaluate the electrostatic effects of the charge placed at these three positions. In this study, we made a series of mutations in three such residues in eukaryotic ClC Cl − channels (K149, G352, and H401 in ClC-0) and studied the functional consequences on the channel properties. It was speculated that the corresponding residues in eukaryotic Cl − channels might play similar roles for the channel functions. Theoretical calculations based on the bacterial ClC structures have identified several residues critical for the Cl − binding energy in the Cl − transport pathway. The recently solved structures of the bacterial ClC proteins have provided a good model to guide the functional experiments for the eukaryotic Cl − channels. The ClC family of Cl − channels and transporters comprises membrane proteins ubiquitously present in species ranging from prokaryotes to mammals. The Journal of General Physiology Rockefeller University Press Submitted: 17 November 2005 Accepted: 13 March 2006 Footnotes Abbreviations used in this paper: HEK, human embryonic kidney MTS, methane thiosulfonate MTSEA, 2-aminoethyl MTS MTSEH, 2-hydroxyethyl MTS MTSES, 2-sulfonatoethyl MTS WT, wild-type. The control of the accessibility to charged MTS reagents and the regulation of the anion permeation support the idea that K149 exerts an electrostatic effect on the channel function, confirming the prediction from computational studies. ![]() Zhang, Xiao-Dong Li, Yong Yu, Wei-Ping Chen, Tsung-Yu ![]() Roles of K149, G352, and H401 in the Channel Functions of ClC-0: Testing the Predictions from Theoretical Calculations Roles of K149, G352, and H401 in the Channel Functions of ClC-0: Testing the Predictions from. ![]()
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