Calcium-ion-transporting activity in two microsomal subfractions from rat pancreatic acini. Modulation by carbamylcholine.
Calcium-ion-transporting activity in two microsomal subfractions from rat pancreatic acini. Modulation by carbamylcholine.
复制标题
大鼠胰腺腺泡的两种微粒体亚组分中的钙离子转运活性。
DOI：
10.1042/bj2190679
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发表时间：
1984
期刊：
影响因子：
--
通讯作者：
R. Dormer
中科院分区：
文献类型：
--
作者：
A. E. Richardson;R. Dormer
通过不连续蔗糖密度梯度离心分离大鼠胰腺腺泡产生两个微粒体亚组分，并通过生化标志物进行表征。密度较高的部分（SF 2）是一个高度纯化的粗面内质网制剂;密度较低的部分（SF 1）是异质的，包含高尔基体，内质网和质膜。在两个组分中均证明了在ATP存在下45 Ca 2+的积累以及用二价阳离子载体A23187处理后45 Ca 2+的快速释放。活性~（45）Ca ~（2+）吸收的最适pH值约为1.5。6.8粗面内质网（SF 2）和大约。7.5关于SF 1初始速率测量用于确定粗面内质网摄取系统对游离Ca 2+的亲和力。表观Km为0.16 +/- 0.06 microM，Vmax。每毫克蛋白质的Ca ~（2+）浓度为21.5 +/-5.6 nmol/min。SF 1的45 Ca 2+摄取对Ca 2+不太敏感，在1-2 μ M游离Ca 2+时发生半数最大摄取。当馏分制备从孤立的腺泡刺激3 μ M-氨甲酰胆碱，45 Ca 2+的摄取增加在粗面内质网。摄取增加是由于Vmax较高。Km无明显变化。SF 1对~（45）Ca ~（2+）的摄取无明显影响。总之，两个不同的非线粒体，ATP依赖的钙摄取系统已被证明在大鼠胰腺腺泡。其中一个位于粗面内质网，但另一个的精确位置尚未确定。我们已经表明，粗面内质网中的Ca 2+转运活性可能在维持静息腺泡细胞胞质游离Ca 2+浓度方面具有重要作用，并且参与酶分泌刺激过程中发生的Ca 2+运动。
Two microsomal subfractions from isolated rat pancreatic acini were produced by centrifugation through a discontinuous sucrose density gradient and characterized by biochemical markers. The denser fraction ( SF2 ) was a highly purified preparation of rough endoplasmic reticulum; the less-dense fraction ( SF1 ) was heterogeneous and contained Golgi, endoplasmic reticulum and plasma membranes. 45Ca2+ accumulation in the presence of ATP and its rapid release after treatment with the bivalent-cation ionophore A23187 were demonstrated in both fractions. The pH optimum for active 45Ca2+ uptake was approx. 6.8 for the rough endoplasmic reticulum ( SF2 ) and approx. 7.5 for SF1 . Initial rate measurements were used to determine the affinity of the rough-endoplasmic-reticulum uptake system for free Ca2+. An apparent Km of 0.16 +/- 0.06 microM and Vmax. of 21.5 +/- 5.6 nmol of Ca2+/min per mg of protein were obtained. 45Ca2+ uptake by SF1 was less sensitive to Ca2+, half-maximal uptake occurring at 1-2 microM-free Ca2+. When fractions were prepared from isolated acini stimulated with 3 microM-carbamylcholine, 45Ca2+ uptake was increased in the rough endoplasmic reticulum. The increased uptake was due to a higher Vmax. with no significant change in Km. No effect was observed on 45Ca2+ uptake by SF1 . In conclusion, two distinct non-mitochondrial, ATP-dependent calcium-uptake systems have been demonstrated in rat pancreatic acini. One of these is located in the rough endoplasmic reticulum, but the precise location of the other has not been determined. We have shown that the Ca2+-transporting activity in the rough endoplasmic reticulum may have an important role in maintaining the cytosolic free Ca2+ concentration in resting acinar cells and is involved in Ca2+ movements which occur during stimulation of enzyme secretion.