Screening of hepatocyte proteins binding to complete S protein of hepatitis B virus by yeast-two hybrid system.
OBJECTIVE
To investigate the biological function of complete S protein and to look for proteins interacting with complete S protein in hepatocytes.


METHODS
We constructed bait plasmid expressing complete S protein of HBV by cloning the gene of complete S protein into pGBKT7, then the 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 of plasmids from positive (blue) colonies, we underwent sequence analysis by bioinformatics.


RESULTS
Nineteen colonies were selected and sequenced. Among them, five colonies were Homo sapiens solute carrier family 25, member 23 (SLC25A23), one was Homo sapiens calreticulin, one was human serum albumin (ALB) gene, one was Homo sapiens metallothionein 2A, two were Homo sapiens betaine-homocysteine methyltransferase, three were Homo sapiens Na(+) and H(+) coupled amino acid transport system N, one was Homo sapiens CD81 antigen (target of anti-proliferative antibody 1) (CD81), three were Homo sapiens diazepam binding inhibitor, two colonies were new genes with unknown function.


CONCLUSIONS
The yeast-two hybrid system is an effective method for identifying hepatocyte proteins interacting with complete S protein of HBV. The complete S protein may bind to different proteins i.e., its multiple functions in vivo.
Bai
Gui-Qin
GQ
Department of Obstetrics and Gynecology of First Hospital, Xi'an Jiaotong University, Jiankang Road 1, Xi'an 710061, Shaanxi Province, China.
Cheng
Jun
J
Zhang
Shu-Lin
SL
Huang
Yan-Ping
YP
Wang
Lin
L
Liu
Yan
Y
Lin
Shu-Mei
SM
D001483:Base Sequence; D045744:Cell Line, Tumor; D015723:Gene Library; D006515:Hepatitis B virus / Q000378:metabolism*; D022781:Hepatocytes / Q000378:metabolism*; D006801:Humans; D010957:Plasmids; D011506:Proteins / Q000378:metabolism*; D011995:Recombination, Genetic; D017385:Sequence Homology; D020798:Two-Hybrid System Techniques; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*; D015003:Yeasts
D011506:Proteins; D014764:Viral Proteins