TABLE 2.

Effects of the addition of wt and mutated rPCBP2 on PV RNA replication and PV IRES-mediated translation in PCBP-depleted extracts

Final concn of rPCBP2 (nM)Fold stimulation over control reactiona
wt PCBP2mKH1-2mKH2-6mKH3-3mKH3-4
Repl.Trans.Repl.Trans.Repl.Trans.Repl.Trans.Repl.Trans.
3003.58.4NDbND3.04.12.21.21.40.8
6004.511.11.00.73.76.13.01.52.61.1
1,2002.912.51.10.84.57.43.51.93.61.7
  • a Fold stimulation values represent the pixel volume of the [α-32P]CTP-labeled RibPVE2A(MluI) RNA band (normalized for loading and recovery against the ethidium bromide-labeled 18S and 28S rRNA) or of the [35S]methionine-labeled PV structural protein precursor (normalized for loading against the 3CD band) over a buffer control reaction pixel volume as determined by phosphorimager analysis. These data represent values obtained from the analysis of the scans presented in Fig. 8. Variability in the effectiveness of PCBP depletion from extract to extract resulted in the differences observed upon comparison of the fold stimulation values in Tables 1 and 2. This difference is seen when comparing the wt rPCBP2 fold stimulation values from experiments carried out in extracts that were depleted of a majority of endogenous PCBP (this table) to fold stimulation values from extracts which retained higher levels of endogenous PCBP (Table 1). Repl., replication; Trans., translation.

  • b ND, not determined.