Enrichment for RNA molecules that bind a Diels-Alder transition state analog.
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 approximately 10(14) sequences 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.
Morris
K N
KN
Department of Molecular, Cellular and Developmental Biology, University of Colorado, Boulder 80309.
Tarasow
T M
TM
Julin
C M
CM
Simons
S L
SL
Hilvert
D
D
Gold
L
L
D001483:Base Sequence; D002384:Catalysis; D016384:Consensus Sequence; D008969:Molecular Sequence Data; D012313:RNA / Q000737:chemistry*; D013329:Structure-Activity Relationship
D012313:RNA