Nickel2+-Mediated Assembly of an RNA-Amino Acid Complex
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The RNA species SHR1 reacts with biocytin (
-biotinoyl-L-lysine) in the presence of Ni2+ or Pt2+ to produce a metal-bridged complex that migrates more slowly than unreacted RNA in the presence of streptavidin (StrAv) on denaturing polyacrylamide gels. Mapping of reverse transcription pause sites identified G79 as a reactive nucleotide. G79 is near the 3′ end of a 37 nucleotide core motif that is nearly as reactive as SHR1. SHR1 reacts with biocytin in the presence of Pt2+ to yield a product that comigrates with the Ni2+ product but that is much more stable, suggesting that the metal ion used in the reaction is present in the product, possibly linking the RNA to the amino acid. In support of this model, SHR1 shows a strong affinity for Ni2+ in immobilized metal ion chromatography.
-biotinoyl-L-lysine) in the presence of Ni2+ or Pt2+ to produce a metal-bridged complex that migrates more slowly than unreacted RNA in the presence of streptavidin (StrAv) on denaturing polyacrylamide gels. Mapping of reverse transcription pause sites identified G79 as a reactive nucleotide. G79 is near the 3′ end of a 37 nucleotide core motif that is nearly as reactive as SHR1. SHR1 reacts with biocytin in the presence of Pt2+ to yield a product that comigrates with the Ni2+ product but that is much more stable, suggesting that the metal ion used in the reaction is present in the product, possibly linking the RNA to the amino acid. In support of this model, SHR1 shows a strong affinity for Ni2+ in immobilized metal ion chromatography.