THE MINIMUM PORE DIAMETER OF THE M2 P-2'Δ MUTATION IN THE α1 GLYCINE RECEPTOR GIVES FURTHER INSIGHT INTO FACTORS CONTROLLING GLYCINE RECEPTOR ION CHARGE SELECTIVITY
D.J.S. Lee1, A. Keramidas1, A.J. Moorhouse1, K.D. Pierce2, P.R. Schofield2, P.H. Barry1, 1 Dept of Physiology & Pharmacology, University of New South Wales, Sydney 2052, 2 The Garvan Institute of Medical Research, Darlinghurst, Sydney 2010.
The glycine receptor belongs to a family of ligand-gated ion channel receptors. It is pentameric with each subunit having four transmembrane domains (M1-M4), with each M2 domain lining the channel pore (e.g., Karlin and Akabas, 19951). The P-2'Δ mutation is a single amino acid deletion that removes the proline from the M2 domain, which has been shown to decrease the anion/cation permeability ratio, PCl/PNa, from ~25-28 (WT, Wild type) to a low 3.82. Estimates of the minimum pore diameter of the P-2'Δ were done using whole cell bi-ionic potential measurements, where a range of organic anion solutions were applied to the cells at different membrane voltages to determine permeabilities from their current-voltage relationships and reversal potentials. In general, the anion permeability decreased as the anion diameter increased. The permeabilites for each anion were plotted against the smallest anion diameter (since none of these anions were all non-spherical) and fitted to an equation to determine minimum pore diameter. We estimated the minimum pore diameter of this (P-2'Δ) mutant GlyR to be about 6.1Å compared to a WT value of 5.3Å2. This is still smaller than the values for the cation selective mutants1 with minimum pore diameters ranging from 6.5Å (PCl/PNa = 0.3 ) to 9.7Å (PCl/PNa = 0.1 )2. Our new measurements extend the work of Keramidas et al. (2002)2 and showed that for anion selective, as well as cation selective, GlyR mutants that there is an increase in the relative cation/anion permeability with the minimum pore diameter of the channel. This further supports the hypothesis that ion charge selectivity in ligand-gated ion channels is dependent on both electrostatic effects and changes in minimum pore diameter in the selectivity filter region.
(1) Karlin, A, Akabas MH. Neuron. 1995;15:1231-1244.
(2) Keramidas A, Moorhouse AJ, Pierce KD, Scholfield PR, Barry PH. Journal of General Physiology. 2002;119:393-410.
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