Botanisches Institute der Universität, Venusbergweg 22, D-5300, Bonn 1, Federal Republic of Germany.
Planta. 1983 Jan;159(1):84-90. doi: 10.1007/BF00998818.
Endoplasmic reticulum membranes were isolated from roots of garden cress (Lepidium sativum L. cv Krause) using differential and discontinuous sucrose gradient centrifugation. The endoplasmic reticulum fraction was 80% rough endoplasmic reticulum oriented with the cytoplasmic surface directed outward and contaminated with 12% unidentified smooth membranes and 8% mitochondria. Marker enzyme analysis showed that the activity for endoplasmic reticulum was enriched 2.4-fold over total membrane activity while no other organelle activity showed an enrichment. All evidence indicated that the fraction was composed of highly enriched endoplasmic reticulum membranes. Ca(2+) uptake activity was measured using the filter technique described by Gross and Marmé (1978). The results of these experiments showed an ATP-dependent, oxalate-stimulated Ca(2+) uptake into vesicles of the endoplasmic reticulum fraction. The majority of the transport activity was microsomal since specific inhibitors of mitochondrial Ca(2+) transport (ruthenium red, LaCl3 and oligomycin) inhibited the activity by only 25%. Sodium azide showed no inhibition. The transport was likely directly coupled to ATP hydrolysis since there was no inhibition with carbonylcyanidem-chlorophenylhydrazone. The transport activity was specific for ATP showing only 36% and 29% of the activity with inosine diphosphate and guanosine 5'-triphosphate, respectively. The results indicate a Ca(2+) transport function located on the endoplasmic reciculum of garden cress roots.
使用差速和不连续蔗糖梯度离心法从花园水芹（Lepidium sativum L. cv Krause）的根中分离内质网膜。内质网部分为 80%粗面内质网，细胞质面向外定向，污染 12%未鉴定的光滑膜和 8%线粒体。标记酶分析表明，内质网的活性比总膜活性富集了 2.4 倍，而其他任何细胞器的活性都没有富集。所有证据表明，该部分由高度富集的内质网膜组成。使用 Gross 和 Marmé（1978）描述的过滤技术测量 Ca（2+）摄取活性。这些实验的结果表明，ATP 依赖性草酸刺激的 Ca（2+）摄取进入内质网部分的囊泡中。由于线粒体 Ca（2+）转运的特异性抑制剂（钌红、LaCl3 和寡霉素）仅抑制了 25%的活性，因此大部分转运活性是微粒体。由于羰基氰化物氯苯腙没有抑制作用，因此转运可能与 ATP 水解直接偶联。转运活性对 ATP 具有特异性，分别仅显示为肌苷二磷酸和鸟苷 5'-三磷酸的 36%和 29%的活性。结果表明，花园水芹根的内质网上存在 Ca（2+）转运功能。
