Journal of Microbiology and Biotechnology

  • 제18권2호
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  • Pages.328-333
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  • 2008
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  • 1017-7825(pISSN)
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  • 1738-8872(eISSN)
한국미생물·생명공학회 (The Korean Society for Microbiology and Biotechnology)
권호 표지

Selection and Target-Site Mapping of Peptides Inhibiting HCV NS5B Polymerase Using Phage Display

  • Kim, Min-Soo (Department of Bioscience and Biotechnology, Hankuk University of Foreign Studies) ;
  • Park, Chan-Hee (Department of Bioscience and Biotechnology, Hankuk University of Foreign Studies) ;
  • Lee, Jong-Ho (Department of Bioscience and Biotechnology, Hankuk University of Foreign Studies) ;
  • Myung, Hee-Joon (Department of Bioscience and Biotechnology, Hankuk University of Foreign Studies)
  • 발행 : 2008.02.29
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초록

A series of pep tides binding to the HCV NS5B polymerase was selected from phage display peptide libraries. A conserved motif of Ser-Arg-X-Arg/Leu was identified among the selected peptides, and Pep2 (Trp-Ser-Arg-Pro-Arg-Ser-Leu) was chosen for further characterization. The binding of Pep2 to HCV NS5B in vivo was shown by a yeast two-hybrid assay and by subcellular colocalization analysis using immunofluorescence confocal microscopy. The in vitro interaction was also confirmed by GST pulldown assay. The replication of the HCV 1b subgenomic replicon was efficiently inhibited by the presence of the peptide. By using a subtractive biopanning against Pep2, the binding site of the peptide was mapped at the pocket of Pro388 to Pro391 in the thumb subdomain of the polymerase. A yeast two-hybrid analysis using Pro388Ala and Pro391Ala mutants of NS5B confirmed the binding.

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