Construction and characterization of bifunctional TIM-3-EGFP fusion proteins

T cell immunoglobulin mucin-3 (TIM-3) is the first surface molecule to be found that specifically identifies Th1 cells in both mice and humans, and it negatively regulates Th1 responses. TIM-3 protein is a type I membrane protein. Overexpression of membrane proteins is a major problem to overcome in studies of membrane protein structure and function. In this study, a fusion between the gene encoding human TIM-3 and EGFP was successfully constructed and expressed in Escherichia coli. To our knowledge, this is the first time that human TIM-3 has been overexpressed in E. coli. We showed that the TIM-3-EGFP fusion protein mediates the recognition and binding of apoptotic cells. Furthermore, we demonstrated that the interactions of TIM-3-EGFP with apoptotic cells were blocked by TIM-3-Ig fusion proteins. Taken together, these results suggest that the human TIM-3 protein may act as a receptor for apoptotic cells.

► A fusion between the gene encoding human TIM-3 and EGFP was expressed in Escherichia coli.
► This is the first time that human TIM-3 has been overexpressed in E. coli.
► TIM-3-EGFP mediates the recognition and binding of apoptotic cells.
► The interactions of TIM-3-EGFP with apoptotic cells were blocked by TIM-3-Ig.