FIG 3.

Binding of recombinant ELAV/Hu family RBPs to RNA of the ELAV target ewg. (A) SDS-polyacrylamide gel showing Coomassie blue-stained recombinant ELAV family RBPs used for binding assays. For each of the recombinant proteins, 0.5 μg, 1 μg, and 2 μg were loaded. Marker proteins were bovine serum albumin (66 kDa), ovalbumin (45 kDa), and carbonic anhydrase (30 kDa). A bacterial protein copurifying with rFNE (lanes 7 to 9) is indicated by the star next to lane 9. (B) EMSA gel with RNA from the ELAV binding site in ewg (pA2-I). Uniformly ³²P-labeled RNAs (100 pM) were incubated with recombinant proteins (2 nM, 8 nM, 32 nM, 125 nM, and 500 nM) and separated on 4% native polyacrylamide gels. (C) Graphic representation of EMSA data. The percentage of bound RNA [(input RNA − unbound RNA)/input RNA × 100] is plotted against the concentration of recombinant proteins (in molar units) presented as log values.