Template recognition by an RNA-dependent RNA polymerase: identification and characterization of two RNA binding sites on Q beta replicase.
Two different SELEX protocols were used to generate two classes of RNA ligands that bound Q beta replicase with nanomolar equilibrium dissociation constants. One set of RNAs appeared to exist as pseudoknots with conserved loop sequences. These ligands bound Q beta replicase and ribosomal protein S1 with equal affinities, indicating that the RNAs bind the replicase through its S1 subunit. The second class of ligands bound the replicase via a pyrimidine rich region. The two sets of ligands did not compete for binding to Q beta replicase, demonstrating that the two RNA families bind unique sites on the phage polymerase. Because the second class of ligands bound ribosomal protein S1 very poorly, it is likely that the second RNA binding site is located on one of the three remaining replicase subunits. Published sequences of RNAs replicated by Q beta replicase possess similarities to the two classes of RNA ligands, providing a possible solution to the question of template recognition by the phage polymerase.
Brown
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Department of Molecular, Cellular and Developmental Biology, University of Colorado at Boulder 80309-0347, USA.
Gold
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D001482:Base Composition; D001483:Base Sequence; D001665:Binding Sites; D017931:DNA Primers; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009843:Oligoribonucleotides / Q000737:chemistry / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011777:Q beta Replicase / Q000737:chemistry*; D012313:RNA / Q000737:chemistry / Q000378:metabolism*; D015719:Single-Strand Specific DNA and RNA Endonucleases / Q000378:metabolism
D017931:DNA Primers; D009843:Oligoribonucleotides; D012313:RNA; D011777:Q beta Replicase; D015719:Single-Strand Specific DNA and RNA Endonucleases