Intestinal brush border membrane Na+/glucose cotransporter functions in situ as a homotetramer.
Intestinal brush border membrane Na+/glucose cotransporter functions in situ as a homotetramer.
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肠刷状缘膜钠/葡萄糖协同转运蛋白作为同源四聚体在原位发挥作用。
DOI：
10.1073/pnas.87.4.1456
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发表时间：
1990
影响因子：
11.1
通讯作者：
Kempner,ES
中科院分区：
文献类型：
--
作者：
Stevens,BR;Fernandez,A;Hirayama,B;Wright,EM;Kempner,ES
通过辐射灭活测定肠刷状缘膜结合的Na+/葡萄糖协同转运蛋白的功能单元分子大小。用来自 13 MeV (1 eV = 1.602 x 10(-19) J) 线性加速器的高能电子以 0 至 70 Mrad (1 rad = 0.01 Gy) 的剂量照射 (-135 摄氏度) 保存在冷冻保护剂缓冲液中的纯化刷状缘膜囊泡。每次给药后，研究协同转运蛋白的（i）Na（+）依赖性转运活性和（ii）针对克隆兔肠协同转运蛋白的抗体的免疫印迹分析。增加辐射降低了最大Na(+)依赖性协同转运蛋白活性Jmax，但不影响表观Km。运输功能单元的大小为290 +/- 5 kDa。刷状缘膜的免疫印迹分析给出了 Mr 70,000 的单个条带，其强度随着辐射剂量的增加而降低，并给出了 66 +/- 11 kDa 的目标大小。我们得出的结论是，刷状缘膜中肠道Na+/葡萄糖协同转运蛋白的原位活性需要同时存在四个完整、独立、相同的亚基，排列为同四聚体。
The functional unit molecular size of the intestinal brush border membrane-bound Na+/glucose cotransporter was determined by radiation inactivation. Purified brush border membrane vesicles preserved in cryoprotectant buffer were irradiated (-135 degrees C) with high-energy electrons from a 13-MeV (1 eV = 1.602 x 10(-19) J) linear accelerator at doses from 0 to 70 Mrad (1 rad = 0.01 Gy). After each dose, the cotransporter was investigated with respect to (i) Na(+)-dependent transport activity and (ii) immunologic blot analysis with antibodies against the cloned rabbit intestinal cotransporter. Increasing radiation decreased the maximal Na(+)-dependent cotransporter activity Jmax without affecting apparent Km. The size of the transporting functional unit was 290 +/- 5 kDa. Immunologic blot analysis of brush border membranes gave a single band of Mr 70,000, which decreased in intensity with increased radiation dose and gave a target size of 66 +/- 11 kDa. We conclude that activity of the intestinal Na+/glucose cotransporter in situ in the brush border membrane requires the simultaneous presence of four intact, independent, identical subunits arranged as a homotetramer.
膜结合酶的辐射灭活探针：γ-谷氨酰转肽酶、氨肽酶 N 和蔗糖酶。
影响因子：
2.9
作者：
B. Stevens;E. Kempner;E. M. Wright
通讯作者：
E. M. Wright