Overexpression and purification of PreS region of hepatitis B virus antigenic surface protein adr subtype in Escherichia coli.
PreS domain of Hepatitis B virus (HBV) surface antigen is a good candidate for an effective vaccine as it activates both B and T cells besides binding to hepatocytes. This report deals with overexpression and purification of adr subtype of surface antigen that is more prevalent in Pakistan. PreS region, comprising 119 aa preS1 region plus a 55 aa preS2 region plus 11 aa from the N-terminal S region, was inserted in pET21a+ vector, cloned in E. coli DH5alpha cells and expressed in E. coli BL21 codon+ cells. The conditions for over expression were optimized using different concentrations of IPTG (0.01-5 mM), and incubating the cells at different temperatures (23-41 degrees C) for different durations (0-6 h). The cells were grown under the given optimized conditions (0.5 mM IPTG concentration at 37 degrees C for 4 h), lysed by sonication and the protein was purified by ion exchange chromatography. On the average, 24.5 mg of recombinant protein was purified per liter of culture. The purified protein was later lyophilized and stored at -80 degrees.
Abbas
Naaz
N
School of Biological Sciences, University of the Punjab, New Campus, Lahore 54590, Pakistan.
Ahmad
Aftab
A
Shakoori
Abdul Rauf
AR
D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D004279:DNA, Viral / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D005814:Genes, Viral; D042822:Genomic Instability; D006514:Hepatitis B Surface Antigens / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification*; D017325:Hepatitis B Vaccines / Q000235:genetics / Q000302:isolation & purification; D006515:Hepatitis B virus / Q000235:genetics* / Q000276:immunology*; D008969:Molecular Sequence Data; D010154:Pakistan; D010957:Plasmids / Q000235:genetics; D011498:Protein Precursors / Q000235:genetics* / Q000302:isolation & purification*; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification
D017931:DNA Primers; D004279:DNA, Viral; D006514:Hepatitis B Surface Antigens; D017325:Hepatitis B Vaccines; D011498:Protein Precursors; D011994:Recombinant Proteins; C050806:presurface protein 1, hepatitis B surface antigen; C050807:presurface protein 2, hepatitis B surface antigen