Production of chimeric protein to scavenge immune complexes

产生嵌合蛋白以清除免疫复合物

• 批准号: 02557026
• 负责人: FUJITA Teizo
• 金额: $ 9.15万
• 依托单位: Fukushima Medical College
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Developmental Scientific Research (B)
• 财政年份: 1990
• 资助国家: 日本
• 起止时间: 1990 至 1992
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-02557026/
• 关键词: complement receptor; chimeric gene; chimeric protein; PI-anchor; immune complex; 補体レセプタ-; PIアンカ-; PCR; DAF; CR1; 免疫複合体病

This project provides a novel means of scavenging harmful immune complexes in circulation with a chimeric protein anchored on the surface of red blood cells. Complement receptor 1 (CR1) is a transmembrane protein that binds opsonized immune complexes. Decay-accelerating factor (DAF) belongs to a group of membrane proteins anchored to cell surface by, a glycosyl-phosphatidylinositol (GPI)-anchor.A chimeric cDNA, encompassing the functional domain of CR1 and the 3' end of DAF mRNA (encoding GPI-anchored region of mature DAF), was expressed in Chinese hamster ovary (CHO) cells with a spleen focus forming virus-based expression vector (SFFVneo). Positive cells were selected by resistance to an antibiotics neomycin, and then subjected to several rounds of panning with anti-CR1 antibody-coated dishes.Individual subclones containing highly positive cells are now on the way of cloning by limiting dilution. A study will be followed, in which the chimeric protein, purified from the lysates of positive clones, is incorporated into the surface of red blood cells from human and its ability of binding immune complexes is examined in vitro.

该项目提供了一种新颖的方法，可以用固定在红细胞表面的嵌合蛋白循环中清除有害免疫复合物。补体受体1（CR1）是一种结合调子化免疫复合物的跨膜蛋白。衰减加速因子（DAF）属于一组通过糖基 - 磷脂酰肌醇（GPI）landor.a嵌合cDNA固定在细胞表面上的膜蛋白。成熟DAF的GPI锚定区）在中国仓鼠卵巢（CHO）细胞中表达，脾脏焦点形成基于病毒的表达载体（SFFVNEO）。通过对抗生素新霉素的抗性选择阳性细胞，然后用抗CR1抗体涂层菜肴进行几轮平霉素。含有高阳性细胞的个体亚克隆现在通过限制稀释来克隆。将遵循一项研究，其中从正阳性克隆的裂解物中纯化的嵌合蛋白被纳入了人类的红细胞表面，并在体外检查了其结合免疫复合物的能力。

项目成果

Misao MATSUSHITA and Teizo FUJITA: "Activation of the Classical complement Pathway by Mannose-binding Protein in Association With a Novel Cls-like Serine Protease" Journal of Experimental Medicine. 176. 1497-1502 (1992)

Misao MATSUSHITA 和 Teizo FUJITA：“甘露糖结合蛋白与新型 Cls 样丝氨酸蛋白酶结合激活经典补体途径”实验医学杂志。

T.Kashiwai,K.Ichihara,Y.Endo,H.Tamaki,N.Amino & K.Miyai: "Immunological and biological characteristics of recombinant human thyrotropin" Journal of Immunological Methods. 143. 25- (1991)

T.Kashiwai、K.Ichihara、Y.Endo、H.Tamaki、N.Amino

藤田 禎三,川内 康弘: "補体レセプタ-の生体内分布とその生物学的意義" 日本臨床. 48. 372-377 (1990)

Teizo Fujita、Yasuhiro Kawauchi：“补体受体的生物分布及其生物学意义”日本临床研究 48. 372-377 (1990)。

Y.Suzuki,Y.Ogura,O.Otsubo,K.Akagi & T.Fujita: "Selective deficiency of C1s associated with a systemic lupus erythematosus-like syndrome Report of a Case" Arthritis and Rheumatism. 35. (1992)

铃木Y、小仓Y、大坪O、赤木K

Katharine E.Coyne,Sharon e.Hall,E.Scotto Thompson,Miguel A.Arce Taroh Kinoshita,Teizo Fujita,David J.Anstee,Wendell Rosse and Douglas M.Lublin.: "Mapping of Epitopes,Glycosylation Sites, and Compoement Regulatory Domains in Human Decay Accelerating Factor

Katharine E.Coyne、Sharon e.Hall、E.Scotto Thompson、Miguel A.Arce Taroh Kinoshita、Teizo Fujita、David J.Anstee、Wendell Rosse 和 Douglas M.Lublin。：“表位、糖基化位点和成分调节的映射