A novel method for the purification of HIV-1 p24 protein from hybrid Ty virus-like particles (Ty-VLPs).
The self-assembly properties of a protein encoded by the yeast retrotransposon Ty can be exploited to produce large amounts of recombinant, particulate fusion proteins as hybrid Ty virus-like particles (Ty-VLPs). This system has now been adapted to allow the release of the additional protein by incorporation of a protease cleavage site between the yeast carrier protein and the protein of interest. The purification of the additional protein is facilitated by exploiting the ease with which Ty-VLPs can be purified from other yeast cell components due to their particulate nature. We have used this modified system to produce hybrid particles containing the HIV-1 p24 protein downstream of the recognition sequence for the blood coagulation factor Xa. The p24 was released from the particles by proteolytic cleavage and rapidly separated from the residual particulate material using centrifugation and standard chromatography techniques. This procedure has been used to purify milligram quantities of HIV-1 p24 protein that reacts with anti-p24 sera and elicits the production of p24-specific antibodies in experimental animals.
Gilmour
J E
JE
British Bio-technology Ltd., Cowley, UK.
Senior
J M
JM
Burns
N R
NR
Esnouf
M P
MP
Gull
K
K
Kingsman
S M
SM
Kingsman
A J
AJ
Adams
S E
SE
D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D003001:Cloning, Molecular; D004251:DNA Transposable Elements; D004797:Enzyme-Linked Immunosorbent Assay; D015683:Gene Products, gag / Q000276:immunology / Q000302:isolation & purification*; D005822:Genetic Vectors; D015483:HIV Antibodies / Q000096:biosynthesis; D015488:HIV Antigens / Q000276:immunology / Q000302:isolation & purification*; D016655:HIV Core Protein p24; D015497:HIV-1 / Q000032:analysis* / Q000276:immunology; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification; D014758:Viral Core Proteins / Q000276:immunology / Q000302:isolation & purification*
D004251:DNA Transposable Elements; D015683:Gene Products, gag; D015483:HIV Antibodies; D015488:HIV Antigens; D016655:HIV Core Protein p24; D011993:Recombinant Fusion Proteins; D014758:Viral Core Proteins