Rapid and simple purification of human immunodeficiency virus 1 epitope gp41.
In order to develop a reliable and inexpensive serodiagnostic method, part of the transmembrane glycoprotein gene of HIV-1, gp41', (HIV-env 548-646) was cloned into an expression vector, pCT10 with a sequence encoding a hydroxylamine cleavage site and with a part of Lac Z gene (Lac 2": 834 base pairs) as a fusion partner. Overexpression of Lac Z"-gp41' was induced in E. coli and the gp41' fusion protein was purified to homogeneity by centrifugation, hydroxylamine cleavage and an ion-exchange chromatography. Western blot analysis and enzyme-linked immunosorbant assay (ELISA) using the purified gp41 fragment showed high sensitivity and specificity of gp41 as an antigen to detect anti HIV-1 antibodies in testing human sera. These results suggest that this simple and rapid purification method is reliable for obtaining a large quantity of purified gp41'.
Sohn
M J
MJ
Laboratory of Molecular Genetics, Korea Institute of Science and Technology, Daejun.
Cho
S H
SH
Jang
W H
WH
Chong
Y H
YH
Nham
S U
SU
Lee
Y I
YI
D001483:Base Sequence; D003001:Cloning, Molecular; D004279:DNA, Viral / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000235:genetics / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics; D015752:Genes, env; D005822:Genetic Vectors; D015700:HIV Envelope Protein gp41 / Q000235:genetics / Q000302:isolation & purification*; D015658:HIV Infections / Q000175:diagnosis; D015497:HIV-1 / Q000235:genetics / Q000276:immunology*; D006801:Humans; D008969:Molecular Sequence Data; D010957:Plasmids; D014773:Virology / Q000379:methods*
D004279:DNA, Viral; D000939:Epitopes; D015700:HIV Envelope Protein gp41