Characterization of the vitamin D-dependent Ca2+-binding sites in rat intestinal Golgi-enriched membrane fractions.
Characterization of the vitamin D-dependent Ca2+-binding sites in rat intestinal Golgi-enriched membrane fractions.
复制标题
大鼠肠道富含高尔基体的膜组分中维生素 D 依赖性 Ca2+ 结合位点的表征。
DOI：
10.1042/bj2180347
复制
发表时间：
1984
期刊：
影响因子：
--
通讯作者：
Weiser,MM
中科院分区：
文献类型：
--
作者：
Walters,JR;Weiser,MM
大鼠肠高尔基体富集膜组分采取钙2+的维生素D依赖性的过程中，已被证明在15分钟内恢复与静脉注射1，25-二羟基胆钙化醇的维生素D缺乏动物的饱食。本文件报告的研究，这些膜组分的Ca 2+结合位点的特征。在5和400 μ M之间的浓度的Ca 2+的平衡结合显示出显着的减少，在所有浓度的膜来自维生素D缺乏的动物相比，正常对照饮食喂养的动物。结合位点的主要类别对Ca 2+具有相对高的亲和力（KD约为1.5）。3 μ M）。维生素D缺乏没有改变这类位点的亲和力，但将结合Ca 2+的数量从347 +/- 26减少到168 +/- 50 nmol/mg蛋白质（平均值+/- S.D.）。Mg ~（2+）仅在低Ca ~（2+）浓度下抑制结合，这种结合的特征表明两个结合位点之间存在正协同性。等摩尔浓度的Zn ~（2+）、La ~（3+）、Pb ~（2+）和Mn ~（2+）抑制Ca ~（2+）结合超过50%。增加离子强度降低Ca 2+结合不超过一半。结合在pH 7.5时达到最大，在pH 6.3时达到半最大。大量对Ca 2+具有相对高亲和力的结合位点表明，这种结合不太可能与任何特定蛋白质或许多蛋白质上存在的非特异性位点结合，最可能的位点是脂质分子。
Rat intestinal Golgi-enriched membrane fractions take up Ca2+ by a vitamin D-dependent process that has been shown to recover within 15 min of repletion of vitamin D-deficient animals with intravenous 1,25-dihydroxycholecalciferol. The present paper reports studies characterizing the Ca2+-binding sites of these membrane fractions. Equilibrium binding of Ca2+ at concentrations between 5 and 400 microM showed significant decreases at all concentrations in membranes derived from vitamin D-deficient animals when compared with normal control-diet-fed animals. The predominant class of binding sites had a relatively high affinity for Ca2+ (KD approx. 3 microM). Vitamin D-deficiency did not change the affinity of this class of site, but decreased the number from 347 +/- 26 to 168 +/- 50 nmol of Ca2+ bound/mg of protein (means +/- S.D.). Mg2+ inhibited binding only at low Ca2+ concentrations, and the characteristics of this binding suggested positive co-operativity between two binding sites. Equimolar concentrations of Zn2+, La3+, Pb2+ and Mn2+ inhibited Ca2+ binding by over 50%. Increased ionic strength decreased Ca2+ binding by no more than half. Binding was maximal at pH 7.5 and half-maximal at pH 6.3. The large number of binding sites with relatively high affinity for Ca2+ suggests that it is unlikely that this binding is to any specific protein or to non-specific sites present on many proteins, and that the most likely sites are lipid molecules.