Fröhlich O, Jones S C
Department of Physiology, Emory University School of Medicine, Atlanta, Georgia 30322.
J Membr Biol. 1987;98(1):33-42. doi: 10.1007/BF01871043.
Chloride equilibrium exchange was measured in the presence of intracellular and extracellular urea, several different alkylureas and thiourea. Urea half-inhibited Cl exchange at about 2.5 M, but the other, less polar analogs had significantly higher potencies; e.g., butylurea half-inhibited at about 60 mM. Onset and reversal of inhibition occurred within less than 2 sec. The inhibition exhibited no obvious sigmoidal dependence on urea concentration, and at low concentrations dimethylurea was a noncompetitive inhibitor of Cl exchange. However, at higher concentrations the Dixon plots were curved upward and a Hill analysis of the dimethylurea data yielded a Hill coefficient of at least 1.5. When present on only one side of the membrane, the slowly penetrating thiourea inhibited Cl exchange with a higher potency from the outside of the cell. Cl/Br exchange was inhibited less under conditions of self-inhibition of anion exchange than in the absence of self-inhibition. These data indicate that the ureas inactivate the anion transporter by a reversible denaturation process, and that the function of the anion transport mechanism may be more sensitive to small perturbations of protein structure than are spectroscopically derived structural parameters.
在细胞内和细胞外存在尿素、几种不同的烷基脲和硫脲的情况下，测定了氯离子平衡交换。尿素在约2.5 M时对氯离子交换有半数抑制作用，但其他极性较小的类似物效力明显更高；例如，丁基脲在约60 mM时产生半数抑制作用。抑制作用的开始和逆转在不到2秒内发生。抑制作用对尿素浓度没有明显的S形依赖性，在低浓度时，二甲基脲是氯离子交换的非竞争性抑制剂。然而，在较高浓度下，迪克森图向上弯曲，对二甲基脲数据进行希尔分析得出的希尔系数至少为1.5。当硫脲仅存在于膜的一侧时，其缓慢渗透，从细胞外侧抑制氯离子交换的效力更高。在阴离子交换自我抑制的条件下，氯/溴交换受到的抑制比不存在自我抑制时要小。这些数据表明，尿素通过可逆的变性过程使阴离子转运体失活，并且阴离子转运机制的功能可能比光谱学得出的结构参数对蛋白质结构的微小扰动更敏感。
