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. 2009 Mar 4;83(10):5137–5147. doi: 10.1128/JVI.02179-08

FIG. 5.

FIG. 5.

CKB enhances NS3-4A helicase and HCV replicase activities. (A) In vitro RNA helicase activity of NS3-4A or NS3 was determined by detecting unwound single-strand RNA (ss) derived from the partially dsRNA substrate (ds). Band intensities corresponding to unwound products and those to dsRNA substrates were determined by ImageQuant 5.2 (Molecular Dynamics), and the ssRNA/dsRNA ratios were calculated. The results are representative of three similar experiments. (B) In vitro DNA helicase activity of NS3-4A or NS3 was analyzed by using a commercially available kit. The data represent averages and standard deviations (n = 3). *, P < 0.05 against the value without supplementation of CKB and ATP. (C) The in vitro HCV protease activity of NS3-4A or NS3 in the presence or absence of CKB was analyzed. Error bars represent standard deviations (n = 3). (D) Replicase activity in permeabilized replicon cells. The upper panel shows the activity for synthesis of HCV subgenomic RNA in the digitonin-permeabilized SGR-JFH1 cells with or without supplementation of CKB was measured. The middle panel shows results for SGR-JFH1 or Huh-7 cells that were transfected with siCKB-2 or siCont and permeabilized at 72 h posttransfection. The permeabilized cells with or without supplementation of CKB were subjected to the replicase assay. The lower panel shows the immunoblotting results for whole-cell lysates of siRNA-transfected cells.