title: [Expression of osteopontin RGD peptide in E. coli and its biological activities].
abstract: OBJECTIVE
  The six copies of RGD sequences present in osteopontin were expressed in the E. coli, and the biological activity of the purified products was studied.
  
  
  METHODS
  cDNA fragments containing six copies of RGD sequences were subcloned into prokaryotic expression vector pGEX-3X including GST coding sequence to construct pGEX-3X-RGD plasmids. E. coli DH5alpha transformed by pGEX-3X-RGD(6) plasmid was induced by different IPTG concentrations for different times to identify the optimal induction condition. Induced GST-RGD fusion protein was purified via GST-Sepharose 4B affinity resin.
  
  
  RESULTS
  GST-RGD fusion proteins containing six copies of RGD sequences could be successfully induced and were mainly located in inclusion bodies. After being denatured and dialyzed, renatured fusion proteins were purified via GST-Sepharose 4B affinity resin. GST-RGD(6) fusion protein could specifically inhibit adhesion and migration of VSMC stimulated by osteopontin, which could be considered as a basis on producing small peptides containing RGD sequences for inhibition of VSMC adhesion and migration.
  
  
  CONCLUSIONS
  The six copies of osteopontin RGD peptide were successfully expressed in E. coli DH5alpha. The purified GST-RGD fusion protein could inhibit the adhesion and migration of VSMC stimulated by osteopontin.
authors.affiliation: Department of Biochemistry and Molecular Biology, Institute of Basic Medicine, Hebei Medical University, Shijiazhuang 050017, China.
authors.forename: Hong-Yu
authors.initials: HY
authors.lastname: Wu
authors.forename: Jin-Kun
authors.initials: JK
authors.lastname: Wen
authors.forename: Mei
authors.initials: M
authors.lastname: Han
mesh_terms: D000818:Animals; D002448:Cell Adhesion / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects*; D009842:Oligopeptides / Q000235:genetics / Q000378:metabolism*; D053495:Osteopontin / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*
chemical_list: D009842:Oligopeptides; D011993:Recombinant Fusion Proteins; D053495:Osteopontin; C047981:arginyl-glycyl-aspartic acid; D005982:Glutathione Transferase