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. Author manuscript; available in PMC: 2016 Apr 1.
Published in final edited form as: J Biomol NMR. 2015 Jan 29;61(0):209–226. doi: 10.1007/s10858-015-9898-7

Fig. 6. Functional characterization of reductively methylated ClC-ec1.

Fig. 6

a Schematic outline of the Cl- flux assay. ClC-ec1 is reconstituted into liposomes at high [KCl], and then the extravesicular solution is exchanged for a low-[KCl] solution (replacing Cl- with the impermeant isethionate, “X-”). Bulk Cl- efflux through ClC-ec1 is initiated by valinomycin (a K+ ionophore) and CCCP (a H+ ionophore) to dissipate the electrical potential. Extravesicular [Cl-] is monitored using a Ag/AgCl electrode.

b Representative raw data traces for WT ClC-ec1 (red) and control liposomes (containing no protein) (black dashes). At the end of each experiment, the detergent Triton X-100 was added to lyse the liposomes and release all of the Cl-.

c Summary data for reductively methylated (RM) proteins studied here. The relative turnover rates (normalized to WT unlabeled ClC-ec1) were calculated from the initial slope (Δ[Cl-]/Δt) after addition of Vln/CCCP, and averages ± SEM (n=3 to 6) are reported.