[Expression and analysis of biological activity of wild-type and mutant interleukin-13 in E.coli].
OBJECTIVE
To express recombinant wild-type human interleukin-13 (rhIL-13) and mutant interleukin-13 (rhIL-13') in E.coli BL21 (DE3) and get active proteins through purification and renaturation.


METHODS
IL-13 and IL-13' gene fragments were amplified by PCR and site-directed mutagenesis PCR, respectively and then were inserted into expression vector pET30a(+). Recombinant plasmids were transformed to E.coli BL21 (DE3) and were expressed under IPTG induction. Expressed products were purified through Ni-NTA chromatographic column. The purified proteins were renatured by GSH-GSSG (reduced glutathione, oxidized glutathione) system and dialysis and their bioactivity was detected by MTT colorimetry.


RESULTS
The expressed recombinant proteins existed in the form of inclusion body with relative molecular mass about 17 000. The recombinant proteins with higher purity were obtained after purification. The renatured inclusion bodies were biologically active.


CONCLUSIONS
rhIL-13/rhIL-13' with biological activity have been obtained successfully, which lays the foundation for further study on their function.
Jiang
Xiao-feng
XF
The Center of Experiment Diagnosis, Second Hospital Affiliated to Harbin Medical University, Harbin 150086, China. jiangxiaofeng12359@hotmail.com
Guo
Xiao-ying
XY
Wan
Li-ping
LP
D015153:Blotting, Western; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000166:cytology / Q000235:genetics*; D015870:Gene Expression; D006801:Humans; D002479:Inclusion Bodies; D018793:Interleukin-13 / Q000032:analysis / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D050505:Mutant Proteins / Q000032:analysis / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D017422:Sequence Analysis, DNA
D018793:Interleukin-13; D050505:Mutant Proteins; D011993:Recombinant Fusion Proteins