Taglialatela M, Payne J P, Drewe J A, Brown A M
Department of Molecular Physiology and Biophysics, Baylor College of Medicine, Houston, TX 77030.
Biophys J. 1994 Jan;66(1):179-90. doi: 10.1016/S0006-3495(94)80755-6.
In a chimeric, voltage-dependent K+ channel (CHM), the valine at position 369 and the leucine at position 374 interact within the pore or P-region to regulate ion permeation and block. Here we show that the point mutation, CHM V369L, abolished channel function whereas previous experiments showed that CHM V369 and CHM V369I are functional. Coinjection of "lethal" CHM V369L cRNA with CHM L374V cRNA but not CHM cRNA generated functional heteromultimers. The whole-cell Rb+/K+ conductance ratio was 2.98 +/- 0.43 for CHM L374V and was reduced to 0.87 +/- 0.04 for the coexpressed CHM V369L and CHM L374V subunits. When single-channel currents were recorded, a single class of CHM V369L/CHM L374V heteromultimers was identified. This class was readily distinguishable from CHM L374V homomultimers by K+ conductance, gating, and blockade by internal tetraethylammonium. Coinjection experiments at various RNA ratios suggest that the CHM V369L/CHM L374V heteromultime, assuming it to be a tetramer, was composed of three CHM L374V subunits and one CHM V369L subunit. It appears that in the critical P-region of CHM position 369 may tolerate only one leucine.
在一种嵌合的电压依赖性钾离子通道（CHM）中，第369位的缬氨酸与第374位的亮氨酸在孔道或P区域内相互作用，以调节离子通透和阻断。我们在此表明，点突变CHM V369L使通道功能丧失，而先前的实验表明CHM V369和CHM V369I具有功能。将“致死性”的CHM V369L cRNA与CHM L374V cRNA而非CHM cRNA共注射可产生功能性异源多聚体。CHM L374V的全细胞铷离子/钾离子电导比为2.98±0.43，而共表达的CHM V369L和CHM L374V亚基的该比值降至0.87±0.04。当记录单通道电流时，鉴定出了一类CHM V369L/CHM L374V异源多聚体。通过钾离子电导、门控以及内部四乙铵的阻断作用，这类异源多聚体很容易与CHM L374V同源多聚体区分开来。在不同RNA比例下进行的共注射实验表明，假设CHM V369L/CHM L374V异源多聚体为四聚体，它由三个CHM L374V亚基和一个CHM V369L亚基组成。似乎在CHM的关键P区域，第369位可能仅能容纳一个亮氨酸。
