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(a) Recombinant eIF4E or bovine serum albumin (BSA) was titrated against the FP probe ATTO-465-labelled 2′/3′-O-(2-aminoethylcarbamoyl)-7-methyl-guanosine-5′-diphosphate (EDA-m7GDP-ATTO-465, structure shown inset; 10 nM) and a Kd value with respect to eIF4E binding was calculated. (b) The FP assay is specific for the unique structural feature of the mRNA cap and can distinguish between m7GTP and GTP (eIF4E concentration: 90 nM).
