Negatively charged liposome as a potent inhibitor of post-translation during in vitro synthesis of green fluorescent protein
Negatively charged liposome as a potent inhibitor of post-translation during in vitro synthesis of green fluorescent protein
复制标题
DOI：
10.1016/j.bej.2009.05.002
复制
发表时间：
2009-10-01
影响因子：
3.9
通讯作者：
Kuboi, Ryoichi
中科院分区：
文献类型：
--
作者：
Bui, Huong Thi;Umakoshi, Hiroshi;Kuboi, Ryoichi
利用无细胞翻译系统研究了带负电荷的脂质体对报告蛋白绿色荧光蛋白（UP）体外合成的影响。用和不用由1-棕榈酰-2-油酰-sn-甘油-3-磷脂酰胆碱（POPC）和1-棕榈酰-2-油酰-sn-甘油-3-磷酸甘油（POPG）制备的带负电荷的脂质体表达GFP，导致在POPC/POPG以大于30%POPG存在下GFP荧光降低至约60%，这取决于其浓度。然而，如通过SDS-PAGE分析的，合成的GFP产物的量在有和没有POPC/POPG脂质体的情况下没有变化。超滤操作的结果表明，脂质体不仅与合成的多肽GFP相互作用，而且与折叠的合成GFP（成熟GFP）相互作用。脂质体还抑制未折叠的GFP重折叠至其天然状态。以上结果表明，POPC/POPG可以抑制GFP在基因表达产物翻译后过程中的折叠。(C)2009爱思唯尔有限公司版权所有。
The effect of negatively charged liposome on in vitro synthesis of a reporter protein, green fluorescent protein (UP), was investigated using a cell-free translation system. GFP was expressed with and without negatively charged liposome prepared with 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylchorine (POPC) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol (POPG), resulting in the GFP fluorescence being reduced to approximately 60% in the presence of POPC/POPG at more than 30% POPG, depending oil its concentration. However, the amount of synthesized GFP products, as analyzed by SDS-PAGE, did not change with and without the POPC/POPG liposome. The results of the ultrafiltration operation indicate that the liposome interacts not only with synthesized polypeptide GFP but also with folded synthesized GFP (mature GFP). Liposome also inhibited refolding of unfolded GFP to its native state. The above results show that the POPC/POPG could inhibit the folding of GFP in the post-translational process of the gene expression product. (C) 2009 Elsevier B.V. All rights reserved.