Yeast two-hybrid system screening liver cell proteins interacting with the novel protein encoded by the 2.2 kb singly spliced variant of hepatitis B virus genome

VernacularTitle:酵母双杂交筛选与单剪接型2.2 kb乙型肝炎病毒剪接特异性新蛋白相互作用的肝细胞蛋白
Author: Wannan CHEN ; Jinyan CHEN ; Qingling HUANG ; Jianyin LIN ; Xu LIN
Publication Type:Journal Article
Keywords: Hepatitis B virus; RNA splicing; Yeast two-hybrid
From: Chinese Journal of Microbiology and Immunology 2010;30(3):228-233
CountryChina
Language:Chinese
Abstract: Objective To screen the liver cell proteins interacting with the novel protein encoded by the 2.2 kb singly spliced variant of hepatitis B virus genome. Methods The splicing-specific gene TPss generated by the 2.2 kb singly spliced variant of HBV genome was amplified by PCR and cloned into the bait vector pGBKT7. After exclusion of self-activatian capacities of TPss protein, a two-hybrid library screening was performed using a pre-transformed human liver cDNA library to screen the liver cell proteins interacting with TPss. Mammalian two-hybrid assay was also done to further confirm the interactions between the bait and prey proteins in Huh7 and HepG2 hepatacytes. Results TPss gene with the size of 336 bp was successfully amplified and cloned, and the TPss protein expressed well in AHI09 yeast cells. Four liver cell proteins interacting with TPss, i. e. , cathepsin B, epoxide hydrolase 1, cathepsin D and fibrinogen gamma chain, were screened by yeast two-hybrid assay and further confirmed by mammalian two-hybrid assay. Conclusion TPss could interact with liver cell proteins.