C-terminus determinants for Mg2+ and polyamine block of the inward rectifier K+ channel IRK1.
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.
Taglialatela
M
M
Rammelkamp Center for Research, Case Western University, Cleveland, OH 44109-1998, USA.
Ficker
E
E
Wible
B A
BA
Brown
A M
AM
D000595:Amino Acid Sequence; D000818:Animals; D008274:Magnesium / Q000494:pharmacology*; D008969:Molecular Sequence Data; D009154:Mutation; D009865:Oocytes / Q000378:metabolism; D011073:Polyamines / Q000494:pharmacology*; D026902:Potassium Channel Blockers; D015221:Potassium Channels / Q000235:genetics / Q000378:metabolism*; D024661:Potassium Channels, Inwardly Rectifying; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D014981:Xenopus
D011073:Polyamines; D026902:Potassium Channel Blockers; D015221:Potassium Channels; D024661:Potassium Channels, Inwardly Rectifying; D008274:Magnesium