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KMID : 0545120010110020234
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Journal of Microbiology and Biotechnology
2001 Volume.11 No. 2 p.234 ~ p.241
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Expression of the Functional Recombinant Interleukin-16 in E. coli and Mammalian Cell Lines
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Kim, Seon Young
Lee, Chang Hun/Kim, Kyung Joo/Kim, Yeon Soo
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Abstract
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The C-terminal 393bp region of the human interleukin-16 (IL-16) gene was cloned and expressed in E. coli along with mammalian cell lines. Recombinant IL-16 expressed from E. coli was 22kDa on SDS-PAGE and showed 260% of chemoattractant activity at a concentration of 0.1㎍/ml. HeLa, COS, and Neuro-2a cells were transduced by recombinant retrovirus vector pLNC/IL-16/IRES/TK and the intracellular and secreted amounts of IL-16 produced by HeLa/IL-16/TK, COS/IL-16/TK, and Neuro-2a/IL-16/TK cells were determined by enzyme-linked immunosorbent assay (ELISA). HeLa/IL16/TK (1×10^5) and COS/IL-16/TK (l×10^5) cells secreted 36.1 and 13.3ng of IL-16 for 48h, respectively. Forty-nine ng and 86.4ng of IL-16 remained in the cell lysates of HeLa/IL-16/TK and COS/IL-16/TK. Intracellular and secreted amounts of IL-16 from Neuro-2a/IL-16/TK (5×10^5) cells during 24h cultivation were 50ng and 3.3ng, respectively. Also, HeLa and COS cells were stably transfected with mammalian expression vector pCRIII/IL-16. Both culture media and cell lysates prepared from HeLa/IL- 16 cells and COS/IL-16 cells showed chemoattractant activity ranging from 190% to 460% as compared to the control experiment. Expression of the herpes simplex virus thymidine kinase (HSV-tk) gene in pLNC/IL-16/IRES/TK bicistronic retroviral expression vector was verified by performing a ganciclovir (GCV) sensitivity assay. Finally, IL-16 repressed Tat-transactivated human immunodeficiency virus type 1 long terminal repeat (HIV-1 LTR) promoter activity.
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