| Autor: |
Mitola S; Department of Molecular and Translational Medicine, University of Brescia, Viale Europa 11, 25123 Brescia, Italy., Ravelli C; Department of Molecular and Translational Medicine, University of Brescia, Viale Europa 11, 25123 Brescia, Italy., Corsini M; Department of Molecular and Translational Medicine, University of Brescia, Viale Europa 11, 25123 Brescia, Italy., Gianoncelli A; Department of Molecular and Translational Medicine, University of Brescia, Viale Europa 11, 25123 Brescia, Italy., Galvagni F; Department of Biotechnology, Chemistry and Pharmacy, University of Siena, 53100 Siena, Italy., Ballmer-Hofer K; Biomolecular Research, Paul Scherrer Institute, 5232 Villigen, Switzerland., Presta M; Department of Molecular and Translational Medicine, University of Brescia, Viale Europa 11, 25123 Brescia, Italy., Grillo E; Department of Molecular and Translational Medicine, University of Brescia, Viale Europa 11, 25123 Brescia, Italy. |
| Abstrakt: |
Gremlin-1 is a secreted cystine-knot protein that acts as an antagonist of bone morphogenetic proteins (BMPs), and as a ligand of heparin and the vascular endothelial growth factor receptor 2 (VEGFR2), thus regulating several physiological and pathological processes, including embryonic development, tissue fibrosis and cancer. Gremlin-1 exerts all these biological activities only in its homodimeric form. Here, we propose a multi-step approach for the expression and purification of homodimeric, fully active, histidine-tagged recombinant gremlin-1, using mammalian HEK293T cells. Ion metal affinity chromatography (IMAC) of crude supernatant followed by heparin-affinity chromatography enables obtaining a highly pure recombinant dimeric gremlin-1 protein, exhibiting both BMP antagonist and potent VEGFR2 agonist activities. |
