title: [Construction of SDF-1P2G54, a specific antagonist of CXCR4].
abstract: OBJECTIVE
  To obtain a specific antagonist of CXCR4, SDF-1P2G54 by mutating SDF-1 second proline (P) into glycin (G) and removing the α-helix of its C-terminal.
  
  
  METHODS
  SDF-1p2g54 gene amplified by PCR was inserted into the vector pET-30a (+) and transformed into Escherichia coli (E. coli) strain BL21. After IPTG induction of E. coli, the expressed recombinant protein was purified with nickel-affinity chromatography column under denaturing conditions and refolded with gradient dilution and ultra-filtration. The chemotactic effect of SDF-1P2G54 on Jurkat cells and its antagonistic effect against SDF-1 were determined by transwell assay; flow cytometry was used to assay the ability of SDF-1P2G54 to induce calcium influx and CXCR4 internalization in MOLT4 cells.
  
  
  RESULTS
  The recombinant protein SDF-1P2G54 completely lost the functions to activate CXCR4 or to induce transmembrane migration of Jurkat cells and calcium influx in MOLT4 cells, but maintained a high affinity to CXCR4. SDF-1P2G54 effectively inhibited the chemotactic effect of wild-type SDF-1 to Jurkat cells, and induced rapid CXCR4 internalization in MOLT4 cells.
  
  
  CONCLUSIONS
  SDF-1P2G54 is a new antagonist of CXCR4 with a potential value as an effective inhibitor of HIV-1 infection, cancer metastasis or other major diseases.
authors.affiliation: Department of Microbiology, Southern Medical University, Guangzhou, China. sugaryang.2011@ yahoo.cn
authors.forename: Feihua
authors.initials: F
authors.lastname: Yang
authors.forename: Beiguo
authors.initials: B
authors.lastname: Long
authors.forename: Yi
authors.initials: Y
authors.lastname: Tan
authors.forename: Ya
authors.initials: Y
authors.lastname: Gong
authors.forename: Weifeng
authors.initials: W
authors.lastname: Ma
mesh_terms: D002460:Cell Line; D019743:Chemokines, CXC / Q000096:biosynthesis / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006801:Humans; D050505:Mutant Proteins / Q000096:biosynthesis / Q000235:genetics; D019718:Receptors, CXCR4 / Q000037:antagonists & inhibitors*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics
chemical_list: C527276:CXCR4 protein, human; D019743:Chemokines, CXC; D050505:Mutant Proteins; D019718:Receptors, CXCR4; D011994:Recombinant Proteins; C452845:SDF-1P2G