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J. Microbiol. Biotechnol. 2001; 11(2): 234-241

Published online April 28, 2001

Copyright © The Korean Society for Microbiology and Biotechnology.

Expression of the Functional Recombinant Interleukin - 16 in E. coli and Mammalian Cell Lines

Seon-Young Kim , Chang-Hun Lee 1, Kyung-Joo Kim 1 and Yeon-Soo Kim 1

Laboratory of Cell Biology Korea Research Institute of Bioscience and Biotechnology 52 Eoundong Yusong Taejon 305-333 Korea, 1Laboratory of Biochemical Genetics National Heart Lung and Blood Institute National Institutes of Health 10 Center Drive Bethesda MD 20892-1654 U.S.A.

Abstract

The C-terminal 393 bp 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 22 kDa on SDS-PAGE and showed 260% of chemoattractant activity at a concentration of $0.1\;{\mu}g/ml$. 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/IL-16/TK $(1{\times}10^5)$ and COS/IL-16/TK $(1{\times}10^5)$ cells secreted 36.1 and 13.3 ng of IL-16 for 48 h, respectively. Forty-nine ng and 86.4 ng 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 $(5{\times}10^5)$ cells during 24 h cultivation were 50 ng and 3.3 ng, respectively. Also, HeLa and COS cells wee 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 (HSV0tk) gene in pLNC/IL-16/ IRES/TK bicistronic retroviral expression vector was verified by performing a genciclovir (GCV) sensitivity assay. Finally, IL-16 repressed Tat-transactivated human immunodeficiency virus type 1 long terminal repeat (HIV-1 LTR) promoter activity.

Keywords: Interleukin-16, E.coli, mammaliancells, retrovirusproducercells, lymphocytemigrationactivity, HIV-1LTRsuppression