重金属离子对猪红细胞膜Ca~(2+)-ATP酶的作用

猪红细胞膜Ca~(2+)-ATP酶是一种钙调蛋白(CaM)依赖酶,其活力又依赖巯基的完整性。实验应用Ca~(2+)-ATP酶这一模型体系观察到重金属离子,Pb~(2+)、Cd~(2+)和Hg~(2+)都能替代Ca~(2+),激活CaM,从而激活Ca~(2+)-ATP酶;其最大刺激活力分别为85％、80％和30％,半刺激浓度分别为32、27和0.7μmol/L。当三种重金属离子的浓度增加时,则与Ca~(2+)-ATP酶的巯基结合,抑制酶的活力,Pb2~(2+)、Cd~(2+)和Hg~(2+)的半抑制浓度分别为370、440和2μmol/L。抑制作用为渐进性过程,而刺激作用为即时效应。抑制作用可为巯基化物,特别是二巯基化物所逆转。研究结果提示,CaM可能是重金属中毒最初作用的靶分子,而重金属中毒不仅使CaM“开关”失灵,还可能导致细胞内Ca~(2+)的调节全面失控。

EFFECTS OF HEAVY METAL CATIONS ON Ca~(2+)-ATPase FROM PIG ERYTHROCYTES

Using calcium pump ATPase, a calmodulin (CaM)-dependent and SH-group-indispensable enzyme as a model system, we found that Pb2+,Cd^and Hg2+effectively substituted for Ca2+ to interact with CaM and stimulated the enzyme. The relative extents otvstimulation by Ca2+, Pb2+,Cd2+- and Hg2+ were 100, 85, 80 and 30, the concentrations (μmol/L) giving half-maximum stimulation being 33, 32, 27 and 0.7 respectively .The effect of these cations on Ca2+-ATPase is biphasic: Stimulatory at low concentration and inhibitory at high concentration. The concentrations (μmol/L) of Pb2+, Cd2+ and Hg2+,for half-maximum inhibition were 370, 400 and 2, repectively.Inhibition, of Ca2+-ATPase activity by these cations varied with a concomitant decrease in basal activity, indicating that the inhibition is mainly due to the binding of these cations to Ca2+-ATPase.The inhibitions could be reversed by thiols. Dithiothreitol and dimercaptopropanol completely restored Ca2+-ATPase activity, whereas glutathionine and cystein were less effective.The action of thiols is compatible with the notion that thiols restore the activity by chelating the heavy metal cations on Ca2+-^ATPase.These findings suggest that the stimulation is due to the occupation of Ca2+ binding site of CaM by the heavy metal cations, and the inhibition is caused by the interaction of these cations with Ca2+-ATPase.An elevated level of heavy metal cations in the cell may. chronically activate (via CaM) or inhibit Ca2+-ATP-ase, thus upsetting the normal regulation of Ca2+ signal and causing disturbances to many vital cell functions.