Prokaryotic expression and identification of human soluble DC-SIGN

Objective To express the soluble form of human dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (sDC-SIGN) without any tag protein in E. coli. Methods The target sequence in pGM-DC-SIGN plasmid that contains human DC-SIGN cDNA was amplified by PCR, inserted into prokaryotic expression vector pET17b and identified by restriction mapping and sequencing. The recombinant expression vector was transformed into E. coli BL21 (DE3) cells. The expressed product was purified by anti-DC-SIGN mAb-Sepharose 4B Affinity Chromatography and identified by SDS-PAGE and Western blot assay. Results The DNA fragment of 1 300 bp, which encode the extracellular region of human DC-SIGN, was amplified from pGM-DC-SIGN plasmid and the recombinant expression vector pET-17b-sDC-SIGN was constructed and confirmed by restriction maps and sequencing. The component of Mr 38 000 in the purified recombinant product was detected by SDS-PAGE and could be recognized by anti-DC-SIGN antibodies in Western blot. Conclusion The recombinant human sDC-SIGN protein was expressed without any tag and purified. This work lays the foundation for further research on functions of sDC-SIGN.