Duggan P F
J Biol Chem. 1977 Mar 10;252(5):1620-7.
The effects of monovalent cations on calcium uptake by fragmented sarcoplasmic reticulum have been clarified. Homogenization of muscle tissue in salt-containing solutions leads to contamination of this subcellular fraction with actomyosin and mitochondrial membranes. When, in addition, inorganic cations are contributed by the microsomal suspension and in association with nucleotide triphosphate substrates there is an apparent inhibition of the calcium transport system by potassium and other cations. However, when purified preparations were obtained after homogenization in sucrose medium followed by centrifugation on a sucrose density gradient in a zonal rotor, calcium uptake and the associated adenosine triphosphatase activity were considerably activated by potassium and other univalent cations. When plotted against the log of the free calcium concentration there was only a slight increase in calcium uptake and ATPase activity in the absence of potassium ions but sigmoid-shaped curves were obtained in 100 mM K+ with half-maximal stimulation occurring at 2 muM Ca2+ for both calcium uptake and ATPase activity. The augmentation in calcium uptake was not due to an ionic strength effect as Tris cation at pH 6.6 was shown to be inactive in this respect. Other monovalent cations were effective in the order K+ greater than Na+ greater than NH4+=Rb+=Cs+ greater than Li+ with half-maximal stimulation in 11 mM K+, 16 mM Na+, 25 mM NH4+, Rb+, and Cs+ and in 50 mM Li+. There was nos synergistic action between K+ AND Na+ ions and both calcium uptak and associated ATPase were insensitive to ouabain. Thallous ions stimulate many K+-requiring enzymes and at one-tenth the concentration were nearly as effective as K+ ions in promoting calcium uptake. The ratio of Ca2+ ions transported to P1 released remained unchanged at 2 after addition of K+ ions indicating an effect on the rate of calcium uptake rather than an increased efficiency of uptake. In support of this it was found that during the stimulation of calcium uptake by Na+ ions there was a reduction in the steady state concentration of phosphorylated intermediate formed from [gamma-32P]ATP. It is considered that there is a physiological requirement for potassium ions in the relaxation process.
单价阳离子对肌浆网碎片摄取钙的影响已得到阐明。在含盐溶液中对肌肉组织进行匀浆会导致该亚细胞组分被肌动球蛋白和线粒体膜污染。此外，当微粒体悬浮液中存在无机阳离子并与三磷酸核苷酸底物结合时，钾离子和其他阳离子会对钙转运系统产生明显抑制作用。然而，当在蔗糖培养基中匀浆后，通过区带转子在蔗糖密度梯度上离心获得纯化制剂时，钾离子和其他单价阳离子会显著激活钙摄取及相关的腺苷三磷酸酶活性。以游离钙浓度的对数作图时，在无钾离子的情况下钙摄取和ATP酶活性仅略有增加，但在100 mM K⁺中获得了S形曲线，钙摄取和ATP酶活性的半最大刺激浓度均为2 μM Ca²⁺。钙摄取的增加并非由于离子强度效应，因为在pH 6.6时的Tris阳离子在这方面无活性。其他单价阳离子的有效性顺序为K⁺＞Na⁺＞NH₄⁺=Rb⁺=Cs⁺＞Li⁺，半最大刺激浓度分别为11 mM K⁺、16 mM Na⁺、25 mM NH₄⁺、Rb⁺和Cs⁺以及50 mM Li⁺。K⁺和Na⁺离子之间没有协同作用，并且钙摄取及相关的ATP酶对哇巴因不敏感。铊离子能刺激许多需要K⁺的酶，其浓度为K⁺离子的十分之一时，在促进钙摄取方面几乎与K⁺离子一样有效。加入K⁺离子后，转运的Ca²⁺离子与释放的P1的比率保持在2不变，这表明其对钙摄取速率有影响，而非摄取效率提高。支持这一点的是，发现在Na⁺离子刺激钙摄取过程中，由[γ-³²P]ATP形成的磷酸化中间体的稳态浓度降低。人们认为在舒张过程中对钾离子有生理需求。
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