Hicks G R, Raikhel N V
Department of Energy Plant Research Laboratory, Michigan State University, East Lansing 48824-1312.
Plant Cell. 1993 Aug;5(8):983-94. doi: 10.1105/tpc.5.8.983.
We have begun to dissect the import apparatus of higher plants by examining the specific association of nuclear localization sequences (NLSs) with purified plant nuclei. Peptides to the simian virus 40 (SV40) large T antigen NLS and a bipartite NLS of maize were allowed to associate with tobacco and maize nuclei. Wild-type NLSs were found to compete for a single class of low-affinity binding sites having a dissociation constant (Kd) of approximately 200 microM. Peptides to mutant NLSs, which are inefficient in stimulating import, were poor competitors, as were reverse wild-type and non-NLS peptides. The NLS binding site was proteinaceous and resistant to extraction under conditions where pores were still associated. In addition, immunofluorescence and immunoelectron microscopy indicated that binding was at the nuclear envelope. Overall, plant nuclei may be an excellent system to identify components of the import apparatus.
我们已开始通过研究核定位序列（NLSs）与纯化的植物细胞核的特异性结合来剖析高等植物的输入装置。将猿猴病毒40（SV40）大T抗原NLS的肽段和玉米的双分NLS与烟草和玉米细胞核进行结合。发现野生型NLSs竞争一类单一的低亲和力结合位点，其解离常数（Kd）约为200微摩尔。刺激输入效率低下的突变NLSs的肽段是较差的竞争者，反向野生型肽段和非NLS肽段也是如此。NLS结合位点是蛋白质性质的，在孔仍相关的条件下对提取具有抗性。此外，免疫荧光和免疫电子显微镜表明结合发生在核膜处。总体而言，植物细胞核可能是鉴定输入装置成分的极佳系统。
