Screening of hepatocyte proteins binding to F protein of hepatitis C virus by yeast two-hybrid system.
OBJECTIVE
To investigate the biological function of F protein by yeast two-hybrid system.


METHODS
We constructed F protein bait plasmid by cloning the gene of F protein into pGBKT7, then recombinant plasmid DNA was transformed into yeast AH109 (a type). The transformed yeast AH109 was mated with yeast Y187 (alpha type) containing liver cDNA library plasmid in 2XYPDA medium. Diploid yeast was plated on synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) containing X-alpha-gal for selection and screening. After extracting and sequencing plasmids from positive (blue) colonies, we underwent sequence analysis by bioinformatics.


RESULTS
Thirty-six colonies were selected and sequenced. Among them, 11 colonies were zymogen granule protein, 5 colonies were zinc finger protein, 4 colonies were zinc-alpha-2-glycoprotein, 1 colony was sialyltransferase, 1 colony was complement control protein factor I, 1 colony was vitronectin, and 2 colonies were new genes with unknown function.


CONCLUSIONS
The yeast two-hybrid system is an effective method for identifying hepatocyte proteins interacting with F protein of hepatitis C virus. F protein may bind to different proteins.
Huang
Yan-Ping
YP
Department of Pediatrics, the First Hospital of Xi'an Jiaotong University, Shaanxi Province, China. huangyp423@126.com
Cheng
Jun
J
Zhang
Shu-Lin
SL
Wang
Lin
L
Guo
Jiang
J
Liu
Yan
Y
Yang
Yuan
Y
Zhang
Li-Ying
LY
Bai
Gui-Qin
GQ
Gao
Xue-Song
XS
Ji
Dong
D
Lin
Shu-Mei
SM
Shao
Qing
Q
D045744:Cell Line, Tumor; D015723:Gene Library; D016174:Hepacivirus / Q000235:genetics / Q000378:metabolism*; D022781:Hepatocytes / Q000378:metabolism*; D006801:Humans; D011485:Protein Binding; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D020798:Two-Hybrid System Techniques; D014758:Viral Core Proteins / Q000378:metabolism*
D014758:Viral Core Proteins; C433031:hepatitis C protein F, Hepatitis C virus