title: [Protein expression and function of gene 2 transregulated by hepatitis B virus pre-s1 protein and its cloning].
abstract: OBJECTIVE
  To screen proteins in leukocytes interacting with PS1TP2 by yeast-two hybrid and to view their subcellular localization in HepG2 cells.
  
  
  METHODS
  The function and structure of PS1TP2 were studied by bioinformatic analysis. PS1TP2 gene was amplified and cloned into plasmid pET32a (+) and pGBKT7 to construct recombinant expression vectors pET32a (+)-PS1TP2 and pGBKT7-PS1TP2. They were transduced into E. coli Rosetta strain and yeast AH109. The transformed yeast mated with yeast Y187 containing leukocyte cDNA library plasmid in a 2xYPDA medium. Diploid yeast cells were plated on a synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) for selecting twice and then screening. Then a green fluorescent protein (GFP) expression vector pEGFP-C1-PS1TP2 was established, transduced into HepG2, and its subcellular localization was studied by fluorescence microscopy and confocal microscopy.
  
  
  RESULTS
  Bioinformatic analysis showed that the PS1TP2 gene was located at 6q24.1, the protein was unstable and the aliphatic index was very high. After transformation of the E. coli and yeast AH109, the expression protein showed: (1) the molecular weight of the expressed product was about 41000 Da, and (2) PS1TP2 existed within the cells. Diploid yeast cells were plated on the synthetic dropout nutrient medium containing X-a-gal for selecting twice and then screening. Twenty-six colonies from blue colonies were sequenced, pEGFP-C1-PS1TP2 was successfully expressed in the HepG2 cells, and PS1TP2 was located in the cell plasma.
  
  
  CONCLUSIONS
  A prokaryotic expression vector pET32a(+)-PS1TP2 was constructed successfully and the PS1TP2 was successfully expressed in the yeast system. Genes of PS1TP2 interact with leukocyte proteins. These results bring some new clues for studying the biological functions of HBV.
authors.affiliation: Institute of Infectious Diseases, Beijing Ditan Hospital, Beijing 100011, China.
authors.forename: Dan-Qiong
authors.initials: DQ
authors.lastname: Wang
authors.forename: Jiang
authors.initials: J
authors.lastname: Guo
authors.forename: Jun
authors.initials: J
authors.lastname: Cheng
authors.forename: Jian-Kang
authors.initials: JK
authors.lastname: Zhang
authors.forename: Long-Feng
authors.initials: LF
authors.lastname: Zhao
authors.forename: Yuan
authors.initials: Y
authors.lastname: Hong
authors.forename: Li-Ying
authors.initials: LY
authors.lastname: Zhang
mesh_terms: D001483:Base Sequence; D003001:Cloning, Molecular; D004151:Dipeptides; D015870:Gene Expression; D056945:Hep G2 Cells; D006514:Hepatitis B Surface Antigens / Q000378:metabolism; D006801:Humans; D011498:Protein Precursors / Q000378:metabolism; D011506:Proteins / Q000235:genetics* / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
chemical_list: D004151:Dipeptides; D006514:Hepatitis B Surface Antigens; D011498:Protein Precursors; D011506:Proteins; C050806:presurface protein 1, hepatitis B surface antigen; C509811:tryptophan-leucine