Overexpression and purification of human immunodeficiency virus type 1 env derived epitopes in Escherichia coli.
In order to develop a reliable and inexpensive serodiagnostic method, a part of envelope gene of HIV-1, gp120' and gp41' (HIV-1 env a.a. 295-474 and a.a. 556-647) was cloned into a T7 expression vector (pET3d). The fusion protein (gp120'-gp41') was overexpressed in Escherichia coli, then purified to homogeneity by a simple gel filtration chromatography. Western blot analysis and enzyme-linked immunosorbent assay (ELISA) using the purified fusion protein showed a high sensitivity and a specificity for the detection of anti HIV-1 antibodies in testing human plasma. These results suggest that the expression scheme employing a direct expression vector and the rapid purification method are reliable and applicable for obtaining a large quantity of HIV-1 env protein for diagnoses of HIV-1 infections.
Sohn
M J
MJ
Molecular Biomedicine Research Group, Korea Research Institute of Bioscience and Biotechnology, Daejun, South Korea.
Lee
M E
ME
Park
H S
HS
Nham
S U
SU
Lee
Y I
YI
D000163:Acquired Immunodeficiency Syndrome / Q000097:blood / Q000175:diagnosis* / Q000276:immunology; D001483:Base Sequence; D015153:Blotting, Western; D002850:Chromatography, Gel; D003001:Cloning, Molecular; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D000939:Epitopes / Q000096:biosynthesis / Q000302:isolation & purification; D004926:Escherichia coli; D015686:Gene Products, env / Q000096:biosynthesis* / Q000302:isolation & purification; D015752:Genes, env; D015483:HIV Antibodies / Q000097:blood*; D015699:HIV Envelope Protein gp120 / Q000096:biosynthesis* / Q000302:isolation & purification; D015700:HIV Envelope Protein gp41 / Q000096:biosynthesis* / Q000302:isolation & purification; D015497:HIV-1 / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000302:isolation & purification; D012680:Sensitivity and Specificity
D000939:Epitopes; D015686:Gene Products, env; D015483:HIV Antibodies; D015699:HIV Envelope Protein gp120; D015700:HIV Envelope Protein gp41; D009838:Oligodeoxyribonucleotides; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins