Expression of platelet-derived endothelial cell growth factor in Escherichia coli and confirmation of its thymidine phosphorylase activity

Biochemistry. 1992 Dec 8;31(48):12141-6. doi: 10.1021/bi00163a024.


Platelet-derived endothelial cell growth factor (PD-ECGF) has been expressed in Escherichia coli as a fusion protein with glutathione S-transferase (GST). The fusion protein was purified by one-step affinity chromatography on glutathione-agarose beads, and recombinant PD-ECGF was proteolytically cleaved with thrombin from its GST leader peptide to yield pure protein. Recombinant PD-ECGF stimulated [3H]methylthymidine uptake by endothelial cells in vitro; however, we were unable to detect stimulation of cell proliferation under a wide variety of conditions. We confirm that in accord with the recent report that PD-ECGF and human thymidine phosphorylase are products of the same gene [Furukawa, T., Yoshimura, A., Sumizawa, T., Haraguchi, M., & Akiyama, S. I. (1992) Nature 356, 668] recombinant PD-ECGF has thymidine phosphorylase activity comparable to that of E. coli thymidine phosphorylase. Further, E. coli thymidine phosphorylase was able to mimic the activity of recombinant PD-ECGF in the [3H]methylthymidine uptake assay, and it appears that recombinant PD-ECGF's effect on the uptake of thymidine by endothelial cells may be due to modulation of cellular thymidine pools. The mechanism by which PD-ECGF stimulates angiogenesis remains to be elucidated.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Cattle
  • Cell Division
  • Cells, Cultured
  • Chromatography, Affinity
  • Cloning, Molecular
  • DNA, Recombinant
  • Electrophoresis, Polyacrylamide Gel
  • Endothelium, Vascular / cytology
  • Endothelium, Vascular / metabolism
  • Escherichia coli
  • Fermentation
  • Humans
  • Molecular Sequence Data
  • Plasmids
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Solubility
  • Temperature
  • Thymidine / analogs & derivatives
  • Thymidine / metabolism
  • Thymidine Phosphorylase / genetics*
  • Thymidine Phosphorylase / metabolism


  • DNA, Recombinant
  • Recombinant Proteins
  • methylthymidine
  • Thymidine Phosphorylase
  • Thymidine