Production and simple purification of a protein encoded by part of the gag gene of HIV-1 in the Escherichia coli HB101F+ expression system inducible by lactose and isopropyl-beta-D-thiogalactopyranoside
Language English Country Netherlands Media print
Document type Journal Article, Research Support, Non-U.S. Gov't
- MeSH
- Chromatography, Ion Exchange MeSH
- Electrophoresis, Polyacrylamide Gel MeSH
- Enzyme-Linked Immunosorbent Assay MeSH
- Escherichia coli drug effects genetics metabolism MeSH
- Gene Products, gag biosynthesis isolation & purification MeSH
- HIV-1 genetics MeSH
- Isopropyl Thiogalactoside pharmacology MeSH
- Lactose pharmacology MeSH
- Humans MeSH
- Antibodies, Bacterial isolation & purification MeSH
- Gene Expression Regulation, Viral drug effects MeSH
- Recombinant Proteins chemistry isolation & purification MeSH
- Genes, Viral genetics MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- Gene Products, gag MeSH
- Isopropyl Thiogalactoside MeSH
- Lactose MeSH
- Antibodies, Bacterial MeSH
- Recombinant Proteins MeSH
The development of the Escherichia coli expression system, which was prepared by transferring the F' episome from strain 71/18 to a highly to a transformable F- strain HB101, is described. These new HB101 (F+) cells, which produced high levels of lac repressor, were capable of taking up lactose and grew under strict selection conditions. A relatively simple two-step purification of part of a protein (M(r) 27,000) encoded by the gag gene of HIV-1 in this expression system is described. The supernatant prepared by removal of cell debris was precipitated by 30% saturation of ammonium sulphate. The protein spectrum was characterized by gel electrophoresis, immunoblotting and ion-exchange titration curves. Optimum separation was achieved using a strong anion exchanger (Mono Q) at pH 8.0. The purified protein did not cross-react with antibodies to E. coli.
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