Timmermans J A, Bindels R J, Van Os C H
Department of Cell Physiology, University of Nijmegen, The Netherlands.
J Nutr. 1995 Jul;125(7 Suppl):1981S-1986S. doi: 10.1093/jn/125.suppl_7.1981S.
In enterocytes and erythrocytes a calmodulin-stimulated Ca(2+)-ATPase is the main Ca2+ efflux pathway. Previous studies have shown that in enterocytes this Ca(2+)-pumping ATPase could be stimulated by vitamin D-dependent Ca(2+)-binding protein, calbindin-D9k, in ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA)-free solutions. In contrast, a similar stimulatory effect of calbindin-D9K was not observed in erythrocytes. We reinvestigated the effects of calbindin, parvalbumin and calmodulin on active Ca2+ uptake in membrane vesicles derived from porcine erythrocytes and from rat duodenum. In EGTA-containing solutions, neither calbindin-D28k nor parvalbumin influenced the rate of ATP-dependent Ca2+ uptake in red blood cell-derived vesicles. However, when EGTA-free solutions were used, calbindin D28k and parvalbumin significantly increased ATP-dependent Ca2+ uptake in erythrocyte as well as in enterocyte-derived membrane vesicles. In contrast, calmodulin significantly increased active Ca2+ uptake in erythrocyte vesicles in the absence as well as in the presence of EGTA. In addition, ATP-dependent Ca2+ uptake in the presence of 0.2 microM calmodulin was further increased by parvalbumin in the absence but not in the presence of EGTA. This observation precludes that parvalbumin and calbindin stimulate the plasma membrane Ca(2+)-ATPase by occupying the calmodulin binding site. Our results support the theoretical notion that calbindin and parvalbumin stimulate the Ca(2+)-starved pump by increasing the free Ca2+ in the immediate vicinity of the Ca2+ pump sites.
在肠上皮细胞和红细胞中，钙调蛋白刺激的Ca(2+)-ATP酶是主要的Ca2+外流途径。先前的研究表明，在肠上皮细胞中，这种Ca(2+)泵ATP酶在不含乙二醇双(β-氨基乙醚)-N,N,N',N'-四乙酸(EGTA)的溶液中可被维生素D依赖性Ca(2+)结合蛋白钙结合蛋白-D9k刺激。相比之下，在红细胞中未观察到钙结合蛋白-D9K的类似刺激作用。我们重新研究了钙结合蛋白、小白蛋白和钙调蛋白对源自猪红细胞和大鼠十二指肠的膜囊泡中Ca2+主动摄取的影响。在含EGTA的溶液中，钙结合蛋白-D28k和小白蛋白均不影响红细胞来源囊泡中ATP依赖性Ca2+摄取的速率。然而，当使用不含EGTA的溶液时，钙结合蛋白D28k和小白蛋白显著增加了红细胞以及肠上皮细胞来源膜囊泡中ATP依赖性Ca2+摄取。相比之下，无论有无EGTA，钙调蛋白均显著增加红细胞囊泡中Ca2+的主动摄取。此外，在不存在EGTA但存在EGTA时，小白蛋白进一步增加了0.2 microM钙调蛋白存在下ATP依赖性Ca2+摄取。这一观察结果排除了小白蛋白和钙结合蛋白通过占据钙调蛋白结合位点来刺激质膜Ca(2+)-ATP酶的可能性。我们的结果支持了这样一种理论观点，即钙结合蛋白和小白蛋白通过增加Ca2+泵位点紧邻区域的游离Ca2+来刺激Ca(2+)饥饿的泵。
