RESUMO
Midkine is a heparin-binding growth factor, which plays important roles in the regulation of cell growth and differentiation. The non-tagged recombinant human midkine (rhMK) is therefore required to facilitate its functional studies of this important growth factor. In the present work, rhMK was expressed in Escherichia coli (E. coli) BL21 (DE3). The expression of midkine was efficiently induced by isopropyl-beta-D-thiogalactopyranoside (IPTG). After sonication, midkine was recovered in an insoluble form, and was dissolved in guanidine hydrochloride buffer. Renaturation of the denatured protein was carried out in the defined protein refolding buffer, and the refolded protein was purified using S-Sepharose ion-exchange chromatography. The final preparation of the rhMK was greater than 98% pure as measured by sodium dodecylsulfate-polyacrylamid gel electrophoresis (SDS-PAGE) and reverse phase high performance liquid chromatography (RP-HPLC). The purified rhMK enhanced the proliferation of NIH3T3 cells.
Assuntos
Animais , Humanos , Camundongos , Sequência de Bases , Proliferação de Células , Citocinas , Genética , Farmacologia , Escherichia coli , Genética , Dados de Sequência Molecular , Células NIH 3T3 , Proteínas RecombinantesRESUMO
Chemokine CXCL9/Mig (monokine induced by IFN-?) belongs to the subfamily of chemotactic cytokines known as CXC-chemokines. In vivo CXCL9 is mainly induced by IFN-? in macrophages and primary glial cells. In vitro, CXCL9 can be secreted by cells such as macrophages, microvascular endothelial cells and neutrophils, in response to the synergy of IFN-? and TLR(toll-like receptor) ligands. CXCL9 is a chemoattractant for activated T lymphocytes, tumor-infiltrating T-lymphocytes, but not for neutrophils or monocytes. The receptor specific for CXCL9 is CXCR3, a G protein-coupled protein which has seven transmembrane domain. The structure and the chemical characterization of CXCL9, as well as its effects on autoimmune deseases, allograft rejection, cancer therapy were reviewed.