Krautwald S, Baccarini M
Department of Immunbiology, Fraunhofer Institut for Toxicology and Molecular Biology, Hannover, Germany.
Biochem Biophys Res Commun. 1993 Apr 30;192(2):720-7. doi: 10.1006/bbrc.1993.1474.
CSF-1 is a dimeric peptide growth factor, stabilized by disulfide bonds. We expressed mouse CSF-1 in bacteria as a fusion protein either with glutathione S-transferase (GST) or with a six histidine tag (His-tag). Large amounts of recombinant material were obtained and purified by a single affinity chromatography step. Purified CSF-1-His-tag monomers efficiently dimerized in vitro, but the presence of variable amounts of GST-moiety in CSF-1 preparations obtained by thrombin cleavage of GST-fusion proteins (thrombin-released CSF-1) interfered with dimerization. However, the thrombin-released CSF-1 monomers possessed agonistic activity, being capable of stimulating tyrosine phosphorylation of the CSF-1 receptor and of an array of cellular proteins in living macrophages and of supporting their growth. These results show that CSF-1 dimerization is not essential for receptor activation in vivo.
集落刺激因子-1（CSF-1）是一种由二硫键稳定的二聚体肽生长因子。我们在细菌中表达小鼠CSF-1，使其作为与谷胱甘肽S-转移酶（GST）或六个组氨酸标签（His-tag）融合的蛋白。通过单一亲和层析步骤获得并纯化了大量重组物质。纯化的CSF-1-His-tag单体在体外有效二聚化，但通过凝血酶切割GST融合蛋白获得的CSF-1制剂（凝血酶释放的CSF-1）中存在可变数量的GST部分会干扰二聚化。然而，凝血酶释放的CSF-1单体具有激动活性，能够刺激CSF-1受体以及活巨噬细胞中一系列细胞蛋白的酪氨酸磷酸化，并支持它们的生长。这些结果表明，CSF-1二聚化对于体内受体激活并非必不可少。
