摘要
应用纤维蛋白单克隆抗体IF 5 3,观察当纤维蛋白的“A”位点与另一纤维蛋白D区域的“a”位点结合后纤维蛋白E区的变化 .纤维蛋白原Aα链经赖氨酰肽链内切酶消化后 ,应用反相HPLC分离纯化 ;通过ELISA法检测单克隆抗体IF 5 3与纤维蛋白原及其衍生物的反应情况 ;应用放射免疫法检测RGD合成肽抑制纤维蛋白单体与IF 5 3反应的情况 .发现IF 5 3能与纤维蛋白原Aα链的一个片段反应 ,该片段经氨基酸序列分析显示为纤维蛋白原Aα链氨基末端 (1~ 2 9) .该抗体能与酸溶解的纤维蛋白单体和可溶性纤维蛋白及XDP反应 ,但不能与酸化纤维蛋白原或GPRP反应 ,因此IF 5 3的抗原决定簇在Aα 2 0~ 2 9,与凝血酶作用于纤维蛋白肽A ,暴露出的聚合位点“A”(Aα17~19)紧邻 .当GPRP存在于纤维蛋白原溶液时 ,经凝血酶作用产生这种纤维蛋白单体不能与IF 5 3反应 .Aα(93~ 99) (ILRGDFS)合成肽部分抑制纤维蛋白单体与IF 5 3的反应 .实验结果提示 ,当纤维蛋白单体相互聚合 ,或纤维蛋白单体与纤维蛋白原聚合时 ,纤维蛋白单体结构会发生变化 ,其中Aα2 0~ 2 9片段成为新抗原暴露于E区表面 ,并且Aα2 0~ 2
Using monoclonal antibody IF 53, the change of E domain of a fribin was observed after its polymerization site “A' binding with site “a' on D domain of another fibrin(ogen). Aα chain of fibrinogen was cleaved by lysylendopeptidase, then fractionated on reversed phase HPLC column. The activity of Fbg and its derivatives against IF 53 were analyzed by kinetic enzyme linked immunosorbent assay (ELISA). The activity of fibrin monomer against IF 53, inhibited by RGD peptide, were analyzed by radiolabel assay. It was found that mAb IF 53 could react with a fraction on Aα chain of fibrinogen. Amino acid sequence analysis showed that the fraction was the N amino\|terminal segment of Aα chain (1—29). This antibody reacted with acid solubilized fibrin monomer, soluble fibrin and cross XDP, but not with acid fibrinogen and GPRP, so the epitope of IF 53 was Aα 20—29 of fibrinogen that was near the “A' polymerization site (Aα 17—19) of fibrinogen. It was also found that this kind of fibrin monomer produced by thrombin did not react with IF 53 when GPRP peptide existed in the solution of fibrinogen. The peptide corresponding to Aα 93—99 (ILRGDFS) segment partially inhibited the binding of fibrin monomer to immobilized IF 53. When fibrin monomer polymerized each other or with fibrinogen, the structure of the fibrin monomer had a change, in which Aα 20—29 as a neo antigen was exposed on the surface of E domain. Aα 20—29 might be next to the cell adhesion domain RGD1.
出处
《中国生物化学与分子生物学报》
CAS
CSCD
北大核心
2004年第2期213-218,共6页
Chinese Journal of Biochemistry and Molecular Biology
基金
辽宁省教育厅科研基金 (No .2 0 12 2 15 9)
辽宁省自然科学基金资助项目 (No 2 0 0 110 2 0 5 1)~~