title: Production and characterization of human soluble CD83 fused with the fragment crystallizable region of human IgG1 in Pichia pastoris.
abstract: The cell surface protein CD83 belongs to the immunoglobulin superfamily and is highly expressed on mature dendritic cells. The soluble form of CD83, sCD83, is a potential immune suppressor. In a previous study, recombinant soluble CD83 was expressed in Escherichia coli, resulting in a lack of functional glycosylation. Although eukaryotic cell systems for producing sCD83 offer the advantages of protein processing, folding, and posttranslational modification, these systems are complicated, expensive, and produce low levels of protein. To obtain more efficient expression of sCD83, we expressed human sCD83 fused with fragment crystallizable region of human IgG1 (hIgG1 Fc) in Pichia pastoris. Under the optimal conditions (time of induction, 48 h; inoculum density (OD600), 80; concentration of methanol, 3.0 %; pH 7.0-8.0; concentration of casamino acid, 5.0 %), the purified human sCD83-hIgG1 Fc (hsCD83-Ig) fusion protein existed as dimers at 25-30 mg/L culture. Treatment with PNGase F showed that purified hsCD83-Ig was modified by N-linked glycosylation. Moreover, the hsCD83-Ig expressed in the P. pastoris system could suppress lymphocyte proliferation in ConA-stimulated and one-way mixed lymphocyte reaction systems. Thus, hsCD83-Ig expressed in P. pastoris is functional and may be used in experimental therapies for graft rejection, graft-versus-host disease, and autoimmune diseases.
authors: Key Laboratory of Transplant Engineering and Immunology, Ministry of Health, Regenerative Medicine Research Center, West China Hospital, Sichuan University, Chengdu, 610041, China.
authors: Yujia
authors: Y
authors: Yuan
authors: Lin
authors: L
authors: Wan
authors: Younan
authors: Y
authors: Chen
authors: Meimei
authors: M
authors: Shi
authors: Chengshi
authors: C
authors: Wang
authors: Jiuming
authors: J
authors: Zhao
authors: Xiaofeng
authors: X
authors: Lu
authors: Hao
authors: H
authors: Wang
authors: Yanrong
authors: Y
authors: Lu
authors: Jingqiu
authors: J
authors: Cheng
mesh_terms: D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism*; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D006031:Glycosylation; D006801:Humans; D007074:Immunoglobulin G / Q000235:genetics / Q000378:metabolism*; D007136:Immunoglobulins / Q000235:genetics / Q000378:metabolism*; D007166:Immunosuppressive Agents / Q000378:metabolism*; D008214:Lymphocytes / Q000187:drug effects; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D010843:Pichia / Q000235:genetics / Q000378:metabolism*; D055503:Protein Multimerization; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D000097802:CD83 Antigen
chemical_list: D015703:Antigens, CD; D007074:Immunoglobulin G; D007136:Immunoglobulins; D007166:Immunosuppressive Agents; D008562:Membrane Glycoproteins; D011993:Recombinant Fusion Proteins; D000097802:CD83 Antigen