Fig. 4. Recombinant GLD-1 binds directly and specifically to the GRE of glp-1 RNA. (A) GST pull down of radiolabeled glp-1 34 nt RNAs by GST-GLD-1. Shown are means of two binding reactions±the variance from a single experiment; similar results were seen in three separate experiments. (B) Competitive displacement of GST GLD-1 from ³²P-labeled SCR (34 nt) with unlabeled wild-type or mutant RNAs. A filter-binding assay was used to detect GLD-1 binding to wild-type ³²P-labeled 34 nt RNA that contains both the GRE and GDE. Increasing concentrations of unlabeled 34 nt RNAs were added to the binding reactions. 100% binding is the amount bound in absence of any competitor. Shown is the mean±s.d. of three to four separate binding reactions.