New affinity resin for purification of cap-binding proteins

Cap binding proteins, which recognize the cap structure present at 5' termini of RNA polymerase H transcripts, have been routinely isolated and purified using affinity resins with mononucleotide cap analogs attached. Here we present a new methodology in which dinucleotide cap analog, m(7) GpppG, has been linked to the EAH-Sepharose. The method is based on derivatization of 2, 3'-cis diol of the second nucleotide within the cap structure, with levulinic acid, and subsequent coupling of resulted acetal through its carboxylic group with aminohexyl-agarose.