Lu Chang-Ming, Yu Jian-Feng, Huang Wei-Da, Zhou Xuan, Zhang Wei-Yan, Xi Hong, Zhang Xue-Guang
Biotechnology Research Institute, Soochow University, Suzhou 215007, China.
Sheng Wu Hua Xue Yu Sheng Wu Wu Li Xue Bao (Shanghai). 2002 Nov;34(6):697-702.
Flt3 ligand (FL) is a hematopoietic growth factor, initiating its in tracellular signaling cascade by binding to counterpart receptor and driving receptor dimerization. The native form of soluble FL in vivo is mainly monomeric. In this study, we constructed a rFL-FL fusion protein cDNA by linking two copies of cDNA encoding the soluble domain of FL in tandem and expressed it in Pichia pastoris. On SDS-polyacrylamide gel electrophoresis, the rFL-FL fusion protein showed a molecular weight of 43 kD, agreeing well with the predicted value. The 43 kD protein was further confirmed by Western blot using polyclonal rabbit anti-human FL antibody. The rFL-FL fusion protein exhibited about 10-fold increment in its activity on colony formation of bone marrow progenitor cells. RFL-FL fusion protein also exerted more potent effect than monomeric FL on extending the survival of starving Raji cells.
Flt3配体（FL）是一种造血生长因子，通过与相应受体结合启动其细胞内信号级联反应并驱动受体二聚化。体内可溶性FL的天然形式主要是单体。在本研究中，我们通过串联连接两个编码FL可溶性结构域的cDNA构建了rFL-FL融合蛋白cDNA，并在毕赤酵母中表达。在SDS聚丙烯酰胺凝胶电泳上，rFL-FL融合蛋白显示分子量为43 kD，与预测值吻合良好。用兔抗人FL多克隆抗体进行的蛋白质印迹进一步证实了该43 kD蛋白。rFL-FL融合蛋白在骨髓祖细胞集落形成活性上表现出约10倍的增加。RFL-FL融合蛋白在延长饥饿Raji细胞存活方面也比单体FL发挥更有效的作用。
