文献类型：期刊文献

英文题名：Recombinant human B7.2 IgV-Iike domain expressed in bacteria maintains its co-stimulatory activity in vitro

作者：Yan, XC; Ma, J; Zheng, J; Lai, BC; Geng, YP; Wang, YL; Si, LS

第一作者：Yan, XC

通讯作者：Si, LS[1]

机构：[1]Xi An Jiao Tong Univ, Coll Med, Sch Med, Inst Immunopathol, Xian 710061, Peoples R China

通讯机构：[1]corresponding author), Xi An Jiao Tong Univ, Coll Med, Sch Med, Inst Immunopathol, Xian 710061, Peoples R China.

年份：2002

卷号：115

期号：7

起止页码：1053-1057

外文期刊名：CHINESE MEDICAL JOURNAL

收录：;WOS:【SCI-EXPANDED(收录号:WOS:000177175200022)】；

语种：英文

外文关键词：human B7.2/CD86; immunoglobulin domain; co-stimulation; T lymphocyte activation; fusion protein

摘要：Objective To investigate which of the two immunoglobulin (Ig)-like domains, the immunoglobulin variable region homologous domain IgV (hB7.2 IgV) and the immunoglobulin constant region homologous domain IgC (hB7.2 IgC) on the human B7.2 molecule contains receptor binding sites, and to evaluate whether the B7.2 protein expressed in bacteria has biological activity in vitro. Methods Three fragments of hB7.2 IgV, hB7.2 IgC and the complete extracellular region of human B7.2 containing both the IgV and IgC domains, hB7.2 Ig (V + C), were amplified by PCR and subcloned into pGEM-TEasy. Three recombinants, pGEX-4T-3-hB7.2 IgV, pGEX-4T-3-hB7.2 IgC and pGEX-4T-3-hB7.2 Ig (V + C), were generated by cloning the fragments into a prokaryote expression plasmid (pGEX4T-3) and transformed into the host strain E. coli DH5alpha. The relevant target fusion proteins consisting of GST and hB7.2 IgV, hB7.2 IgC and hB7.2 Ig (V + C), were identified by SDS-PAGE and Western blotting. With the presence of the first signal imitated by anti-CD3 antibody, T cell activation was observed by exposing purified T lymphocytes to each soluble form of the three bacterially-produced human B7.2 fusion proteins by [H-3]-TdR incorporation. Results Three recombinant fusion proteins of human B7.2, GST-hB7.2 IgV, GST-hB7.2 IgC and GST-hB7.2 Ig (V + C) were produced and detected in inclusion body form from engineered bacteria. With the first signal present, T lymphocytes proliferated when co-stimulated by bacterially-produced either GST-hB7.2 Ig (V + C) or GST-hB7.2 IgV fusion proteins, but not by GST-hB7.2 IgC. Conclusions Functional human B7.2 fusion protein can be produced in bacteria. The IgV-like domain of human B7.2 is sufficient for B7.2 to interact with its counter-receptors and co-stimulate T lymphocytes.