KKXX-motif对VKORC1亚细胞定位的影响

Influence of KKXX-motif on VKORC1 Subcellular Localization

目的研究细胞内质网蛋白贮留信号KKXX-motif,对维生素K环氧化物还原酶复合物亚基1(VKORC1)的亚细胞定位的作用。方法利用点突变试剂盒构建VKORC1的KKXX突变体——VKORC1-SSXX;同时应用pEGFP载体构建VKORC1-SSXX和VKORC1-KKXX的绿色荧光融合蛋白表达质粒。瞬时转染HEK293s细胞,观察野生融合蛋白和突变融合蛋白的表达情况。结果野生型融合蛋白荧光分布在胞浆内,而突变型融合蛋白聚集表达。结论KKXX-motif影响了VKORC1的亚细胞定位。

Objective To determine whether the KKXX-motif can influence the subcellular localization of VKORC1. Methods The mutant VKORC1-SSXX was made by site-directed mutagenesis. The motif KAKRH was substituted by SASRH. The wild and mutant VKORC1 were cloned into pEGFP-C2 vector respectively. Then the constructed plasmids were transiently transfected to HEK293s cells, which were observed under confocal microscope. Results Subcellular fraction revealed that GFP staining diffused in cytoplasmic compartment and nucleus in the cells transfected with control vector, predominantly in the cytoplasm of the cells transfected with GFP-VKORC1. By contrast, the mutant VKORC1-SSXX was expressed in an accumulated pattern in cells. Conclusion KKXX-motif could influence VKORC1 subcellular localization.