ABSTRACT
<p><b>OBJECTIVE</b>To express recombinant human interferon lambda2 in E.coli and to study its antiviral activities.</p><p><b>METHODS</b>According to preferred codons used in E.coli, the highly-expressed human interferon lambda2 gene was designed, synthesized and cloned into expression vector pBV220 and transfected into E.coli DH5alpha. The expressed product was purified by using CM FF and size exclusion chromatography. Its antiviral activities were tested on different cells.</p><p><b>RESULTS</b>The expressed product was calculated about 15% of the total E.coli protein. The purified protein reached about 90% purity. Its specific antiviral activity was about 1.5 x 10(6) IU/mg on WISH/VSV test system. It was shown that the antiviral activity of the product on primates-origin cells seemed to be much higher than that on other non-primates-origin cells, indicating that interferon lambda2 possessed more stringent species specificity as compared with interferon-alpha2b. New interferon lambda2 showed similar anti-HBV activity as interferon-alpha2b.</p><p><b>CONCLUSION</b>Recombinant human interferon lambda2 could be expressed on E.coli. The purified product showed more stringent species specificity and similar anti-HBV activity as compared with interferon-alpha2b.</p>
Subject(s)
Animals , Humans , Antiviral Agents , Pharmacology , Cell Line , Cell Line, Tumor , Cell Proliferation , Chlorocebus aethiops , Dose-Response Relationship, Drug , Electrophoresis, Polyacrylamide Gel , Enzyme-Linked Immunosorbent Assay , Escherichia coli , Genetics , Metabolism , Hepatitis B virus , Interleukins , Genetics , Pharmacology , Microbial Sensitivity Tests , Recombinant Proteins , Pharmacology , Vero CellsABSTRACT
To clone KGF-2 gene, get hKGF-2 protein and detemine its activity. The cNDA of human KGF-2 was isolated from fetal lung by RT-PCR and cloned into pBV220 plasmid. The recombinant pBV220-hKGF-2 plasmid was transformed into E. coli (BL21), induced at 42 degrees C for the expression of hKGF-2. Recombinant human KGF-2 was purified from the ultrasonic-treated BL21 by heparin-Sepharose CL-6B treated column chromatography and cation exchange column chromatography. MTT method was used for the determination of its biological activity. SDS-PAGE showed that rhKGF-2 was expressed in E. coli BL21 as soluble protein of approximately 20kD. The rhKGF-2 protein can stimulate the proliferation of NIH3T3 cells significantly from 1 ng/mL to 10 ng/mL. HKGF-2 cDNA wasclned and highly expressed in E. coli BL21 and the purified rhKGF-2 showed the mitogenic activity on NIH3T3 cells.