Acquired immunodeficiency syndrome(AIDS),caused by human immunodeficiency virus(HIV),has become the most dangerous epidemic in this century.HIV can be spread by drug abuse,contaminated blood,and sex.It can be also infected by mother.In addition,the variation of some related gene in host was another significant factor that can influence the dissemination of HIV and the healing of AIDS sufferer.The recent researches have discovered that the dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin(DC-SIGN) was closely correlated with the infection of HIV-1 because DC-SIGN could bind the envelope glycoprotein gp120 of HIV-1.DC-SIGN is composed of a cytoplasmic domain,a transmembrane domain,and an extracellular domain including a carbohydrate recognition domain(CRD) and neck domain.CRD is the important binding site with HIV.The variety of the repeated sequence of DC-SIGN neck region makes the space structure of DC-SIGN change,and has an effect on the bind between the CRD and HIV gp120 further,then HIV can hardly be combined with DC cell,and finally DC cell can not deliver the relative immunologic information caused by HIV to the CD4+ T cells.Whereas the binding of DC-SIGN and HIV is dependent on the binding site which was located in CRD region,and on the other hand,the binding capability is dependent on the repeated sequence of neck region,so in theory the usage of some specific antibody which can completely combine with the DC-SIGN of DC cells may block up the infection of HIV.Thus in the present study we cloned the coding region fragmen of DC-SIGN from human DCs,and constructed expression system of Bombyx mori baculovirus Bm-BacPAK-DC-SIGN expressing DC-SIGN protein fragmen whose biological activity was measured.It establishes foundation for preparing the soluble fully human scFv against DC-SIGN and exploiting new products for prevention and treatment of human immunodeficiency virus infection.Objective:To clone human coding region fragmen of DC-SIGN,to construct expression system of Bombyx mori baculovirus Bm-BacPAK-DC-SIGN expressing DC-SIGN protein fragmen,and to measure the biological activity of it.Methods:Peripheral blood monocytes(PBMC) were isolated from blood of healthy individuals by Ficoll-Hypaque sedimentation and cultured in complete medium containing rhGM-CSF and rhIL-4 which were added every second day to stimulate them to differentiate into mature DCs.Total RNA was extracted from the human DCs and the coding region fragmen of DC-SIGN was obtained from the total RNA by RT-PCR and PCR using designed primers.The PCR products were determined by agarose gel electrophoresis.The coding region fragmen of DC-SIGN was inserted into pMD18-T vector and identified by enzyme digestion and DNA sequencing.The resulting fragment was digested with BamHⅠand EcoRⅠand gel-purified and ligated into BamHⅠand EcoRⅠ-digested pBacPAK8 to generate pBacPAK8-DC-SIGN.Enzyme digestion showed that the DC-SIGN gene fragmen was correctly inserted into pBacPAK8 vector.The pBacPAK8-DC-SIGN were cotransfected into Bombyx mori cell lines with the linearized Bm-BacPAK6 virus genome DNA.After recombination in the cells and screening by the plaque assay,the recombinated virus Bm-BacPAK-DC-SIGN were obtained and again infected into Bombyx mori cell lines to express DC-SIGN protein fragmen which were detected by SDS-PAGE and Western blot.The biological activity of the expressed DC-SIGN protein fragmen was measured by its combination with HIV-1gp120. Results:①The coding region fragment of DC-SIGN was cloned successfully.②Expression system of Bombyx mori baculovirus Bm-BacPAK-DC-SIGN expressing human DC-SIGN protein fragment was constructed.③Human DC-SIGN protein fragment was expressed and it can bind specifically with HIV-1gp120.Conclusion:Human DC-SIGN protein fragment was successfully expressed in expression system of Bombyx mori baculovirus Bm-BacPAK-DC-SIGN,which showed the natural DC-SIGN protein-like biological activity.It became the work basis of the antibody preparation against it and drug study on prevention and treatment of AIDS.