Enrichment for RNA Molecules that Bind a Diels-Alder Transition State Analog

• Published in: Proceedings of the National Academy of Sciences - PNAS 1994-12, Vol.91 (26), p.13028-13032
• Main Authors: Morris, Kevin N., Tarasow, Theodore M., Julin, Carol M., Simons, Shauna L., Hilvert, Donald, Gold, Larry
• Format: Article
• Language: English
• Subjects: Base Sequence; Catalysis; Chemistry; Consensus Sequence; Conserved sequences; Diels Alder reactions; Elution; Ligands; Molecular Sequence Data; Molecules; Nucleotide sequences; Nucleotides; Polymerase chain reaction; Ribonucleic acid; RNA; RNA - chemistry; Structure-Activity Relationship
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.91.26.13028

RNA molecules that bind a transition state analog for a Diels-Alder reaction (Kd= 0.35 ± 0.05 mM) were isolated from a starting pool of ≈1014sequences by affinity chromatography. After the initial rise and plateau of the amount of RNA that eluted with soluble analog, a step gradient elution was used to further enrich the pool for sequences with higher affinities for the target. To our knowledge, the isolation of RNA molecules that bind either a nonplanar or a hydrophobic ligand has not been reported previously. A conserved nucleotide sequence and secondary structure present in many of the RNA molecules are necessary but not sufficient for binding the analog. No catalysts of the targeted Diels-Alder reaction were found among the binders. The absence of catalysis contrasts with previous successful experiments with antibodies and suggests that other strategies may be needed to identify oligonucleotides with diverse catalytic activities.