L-selectin transmembrane and cytoplasmic domains are monomeric in membranes.
L-selectin transmembrane and cytoplasmic domains are monomeric in membranes.
复制标题
L-选择素跨膜结构域和细胞质结构域在膜中是单体。
DOI：
10.1016/j.bbamem.2011.02.006
复制
发表时间：
2011
期刊：
影响因子：
--
通讯作者：
Li,Renhao
中科院分区：
文献类型：
--
作者：
Srinivasan,Sankaranarayanan;Deng,Wei;Li,Renhao
一种名为CLS的重组蛋白已被生产出来，它对应于人L-选择素的C-末端部分，包含其整个跨膜和胞质结构域(残基Ser473-Arg542)，并表征了其在洗涤剂中的低聚状态。CLS在十二烷基硫酸钠-聚丙烯酰胺凝胶中的迁移速度通常与其分子量两倍的复合体的迁移速度相同。然而，更多的研究表明，这是由于细胞质区域的残基，因为该区域的突变或其缺失显著增加了CLS的电泳率。分析超速离心和荧光共振能量转移研究表明，在十二烷基磷胆碱洗涤剂胶束中重建的CLS是单体。在TOXCAT实验中，将L-选择素的跨膜区作为嵌合蛋白的一部分插入到大肠杆菌的内膜中，几乎没有观察到嵌合蛋白的寡聚。总体而言，这些结果表明，L-选择素的跨膜和胞浆结构域缺乏在膜上自结合的倾向，而不是先前报道的密切相关的P-选择素的跨膜结构域的二聚化。本研究为进一步研究L-选择素及其相关蛋白的相互作用提供了限制条件。
A recombinant protein termed CLS, which corresponds to the C-terminal portion of human L-selectin and contains its entire transmembrane and cytoplasmic domains (residues Ser473-Arg542), has been produced and its oligomeric state in detergents characterized. CLS migrates in the SDS polyacrylamide gel at a pace that is typically expected from a complex twice of its molecular weight. Additional studies revealed, however, that this is due to residues in the cytoplasmic domain, as mutations in this region or its deletion significantly increased the electrophoretic rate of CLS. Analytical ultracentrifugation and fluorescence resonance energy transfer studies indicated that CLS reconstituted in dodecylphosphocholine detergent micelles is monomeric. When the transmembrane domain of L-selectin is inserted into the inner membrane of Escherichia coli as a part of a chimeric protein in the TOXCAT assay, little oligomerization of the chimeric protein is observed. Overall, these results suggest that transmembrane and cytoplasmic domains of L-selectin lack the propensity to self-associate in membranes, in contrast to the previously documented dimerization of the transmembrane domain of closely related P-selectin. This study will provide constraints for future investigations on the interaction of L-selectin and its associating proteins.