CATION-SELECTIVE CHANNELS IN THE VACUOLAR MEMBRANE OF SACCHAROMYCES - DEPENDENCE ON CALCIUM, REDOX STATE, AND VOLTAGE
CATION-SELECTIVE CHANNELS IN THE VACUOLAR MEMBRANE OF SACCHAROMYCES - DEPENDENCE ON CALCIUM, REDOX STATE, AND VOLTAGE
复制标题
DOI：
10.1073/pnas.87.20.7824
复制
发表时间：
1990-10-01
影响因子：
11.1
通讯作者：
SLAYMAN, CL
中科院分区：
文献类型：
--
作者：
BERTL, A;SLAYMAN, CL
利用膜片钳技术研究了酿酒酵母液泡膜的功能表达。在不存在供能底物的情况下，其最显著的特征是具有特征电导率为约的阳离子通道。对于对称的100 mM KCl溶液为120 pS，并且在K+和Na+之间具有很小的选择性（PNa+/PK+= 120 pS）。1)但对阳离子的选择性强于阴离子（PCl-/PK+ < 0.1）。通道门控是电压依赖性的;打开概率Po达到最大值（约0.7)在-80 mV的跨膜电压下（细胞质表面负）并且在更负和更正的电压下均下降（即，+80 mV左右）。时间平均的电流-电压曲线显示强整流，负电流（正电荷从液泡侧流向细胞质侧）远大于正电流。开放概率还强烈依赖于细胞质Ca 2+浓度，但对于普通记录条件，仅在非生理高（≥）时才高ImM）Ca2+。然而，还原剂如二硫苏糖醇和2-巯基乙醇平衡通道，使它们可以被微摩尔细胞质Ca 2+激活。通道被氯胺T不可逆地阻断，氯胺T已知特异性氧化暴露的甲硫氨酸和半胱氨酸残基。
The vacuolar membrane of the yeast Saccharomyces cerevisiae, which is proposed as a system for functional expression of membrane proteins, was examined by patch-clamp techniques. Its most conspicuous feature, in the absence of energizing substrates, is a cation channel with a characteristic conductance of .apprxeq. 120 pS for symmetric 100 mM KCl solutions and with little selectivity between K+ and Na+ (PNa+/PK+ .apprxeq. 1) but strong selectivity for cations over anions (PCl-/PK+ < 0.1). Channel gating is voltage-dependent; open probability, Po, reaches maximum (.apprxeq. 0.7) at a transmembrane voltage of -80 mV (cytoplasmic surface negative) and declines at both more negative and more positive voltages (i.e., to 0 around +80 mV). The time-averaged current-voltage curve shows strong rectification, with negative currents (positive charges flowing from vacuolar side to cytoplasmic side) much larger than positive currents. The open probability also depends strongly on cytoplasmic Ca2+ concentration but, for ordinary recording conditions, is high only at unphysiologically high (.gtoreq. 1 mM) Ca2+. However, reducing agents such as dithiothreitol and 2-mercaptoethanol poise the channels so that they can be activated by micromolar cytoplasmic Ca2+. The channels are blocked irreversibly by chloramine T, which is known to oxidize exposed methionine and cysteine residues specifically.