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EMBO J
1995 Nov 15;1422:5532-41. doi: 10.1002/j.1460-2075.1995.tb00240.x.
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C-terminus determinants for Mg2+ and polyamine block of the inward rectifier K+ channel IRK1.
Taglialatela M
,
Ficker E
,
Wible BA
,
Brown AM
.
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Critical loci for ion conduction in inward rectifier K+ channels are only now being discovered. The C-terminal region of IRK1 plays a crucial role in Mg2+i blockade and single-channel K+ conductance. A negatively charged aspartate in the putative second transmembrane domain (position 172) is essential for time-dependent block by the cytoplasmic polyamines spermine and spermidine. We have now localized the C-terminus effect in IRK1 to a single, negatively charged residue (E224). Mutation of E224 to G, Q and S drastically reduced rectification. Furthermore, the IRK1 E224G mutation decreased block by Mg2+i and spermidine and, like the E224Q mutation, caused a dramatic reduction in the apparent single-channel K+ conductance. The double mutation IRK1 D172N+ E224G was markedly insensitive to spermidine block, displaying an affinity similar to ROMK1. The results are compatible with a model in which the negatively charged residue at position 224, E224, is a major determinant of pore properties in IRK1. By means of a specific interaction with the negatively charged residue at position 172, D172, E224 contributes to the formation of the binding pocket for Mg2+ and polyamines, a characteristic of strong inward rectifiers.
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