Riedel N, Fasold H
Biochem J. 1987 Jan 1;241(1):213-9. doi: 10.1042/bj2410213.
In the preceding paper [Riedel & Fasold (1987) Biochem. J. 241, 203-212] we have described a procedure for the preparation of nuclear-envelope vesicles (NE vesicles) from rat liver nuclei. These vesicles, which are largely free of components of the nuclear interior, were employed in an assay system in vitro to study protein translocation across the NE. We found that nuclear proteins such as histones, high-mobility-group proteins and acidic chromosomal proteins are specifically taken up and accumulated in the NE vesicles, whereas there is little or no affinity for non-nuclear proteins like immunoglobulin, myoglobin and cytochrome c. The kinetics of histone uptake into the NE vesicles are similar to those obtained for whole rat liver nuclei, and comparative studies with non-vesicular NEs prepared by deoxyribonuclease I-treatment (DNAase-NEs) indicate that the NE of the vesicles affects the uptake kinetics and increases the capacity for nuclear proteins. The uptake of histones into NE vesicles, but not the binding to DNAase-NEs, can be stimulated by GTP and GDP. Furthermore, we found that even very large molecules can be entrapped in the vesicles during their preparation. These results indicate that the NE vesicles might provide a useful system in vitro with which to investigate the structures and mechanisms involved in protein translocation across the NE.
在前一篇论文[里德尔和法索尔德（1987年），《生物化学杂志》241卷，203 - 212页]中，我们描述了一种从大鼠肝细胞核制备核膜囊泡（NE囊泡）的方法。这些囊泡基本不含核内成分，被用于体外分析系统以研究蛋白质穿过核膜的转运。我们发现，组蛋白、高迁移率族蛋白和酸性染色体蛋白等核蛋白能被特异性摄取并积累在NE囊泡中，而对于免疫球蛋白、肌红蛋白和细胞色素c等非核蛋白几乎没有或没有亲和力。组蛋白摄取到NE囊泡中的动力学与在整个大鼠肝细胞核中获得的相似，并且与通过脱氧核糖核酸酶I处理制备的非囊泡核膜（DNA酶 - NE）的比较研究表明，囊泡的核膜会影响摄取动力学并增加对核蛋白的容纳能力。GTP和GDP能刺激组蛋白摄取到NE囊泡中，但不能刺激其与DNA酶 - NE的结合。此外，我们发现即使是非常大的分子在囊泡制备过程中也能被包裹在其中。这些结果表明，NE囊泡可能提供了一个有用的体外系统，可用于研究蛋白质穿过核膜所涉及的结构和机制。
