Recombinant human B7-H4 expressed in Escherichia coli inhibits T lymphocyte proliferation and IL-2 secretion in vitro.
AIM: To explore the biofunctions of human B7-H4 generated from prokaryotic system. METHODS: The gene of human B7-H4 extracellular region (IgV-like and IgC-like domains) was obtained by PCR from human cDNA FLJ22418 and then inserted into the prokaryotic expression vector pGEX-5X-3 expressing glutathione s-transferase ( GST) fusion protein. After being identified by restriction enzyme digestion and sequencing, the recombinant vector was transferred into host strain E coli BL21-RIL(DE3). A 47 kDa fusion protein ( GST/hB7-H4) was induced by isopropyl-beta-D-thiogalactopyranoside (IPTG) and purified by standard methods reported in the prokaryotic system. The inhibitory effect of GST/hB7-H4 on proliferation of T cells was observed in vitro by CD3mAb activated T-cell culturing system and [(3)H]-thymidine incorporation assay. The concentrations of interleukin-2 and iterferon-g in the supernatants of T cells were determined by ELISA. RESULTS: We successfully constructed the method for high-level expression and purification of the hB7-H4 extracellular domain as GST fusion protein from E coli. The GST/hB7-H4 fusion protein produced in bacteria had obvious biological activity to inhibit T-lymphocyte proliferation and IL-2 secretion. CONCLUSION: The prokaryote expression system could be used to generate hB7-H4 protein with natural spatial conformations and biological functions, which provided an efficient and economical way for the preparation of this target protein.[1]References
- Recombinant human B7-H4 expressed in Escherichia coli inhibits T lymphocyte proliferation and IL-2 secretion in vitro. Mao, Y.X., Chen, Y.J., Ge, Y., Ma, H.B., Yu, J.F., Wu, H.Y., Hu, Y.M., Wang, Q., Shi, Q., Zhang, X.G. Acta Pharmacol. Sin. (2006) [Pubmed]
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