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. 1994 Jun 25;22(12):2255–2263. doi: 10.1093/nar/22.12.2255

Footprinting RNA-protein complexes following gel retardation assays: application to the R-17-procoat-RNA and tat--TAR interactions.

L Pearson 1, C B Chen 1, R P Gaynor 1, D S Sigman 1
PMCID: PMC523682  PMID: 8036153

Abstract

RNA-protein complexes isolated following a gel retardation assay can be footprinted within the gel matrix using the chemical nuclease activities of 4,7-dimethyl-, 5,6-dimethyl-, and 3,4,7,8-tetramethyl-1,10-phenanthroline-copper. These complexes are more reactive than 1,10-phenanthroline-copper but share its reaction preference for bulges and loops. The interaction of the coat protein of R-17 with its viral RNA target and tat- and tat-derived peptides with HIV TAR RNA have been studied. In both cases, the RNA sequence opposite a 2-3 nucleotide bulge are protected. Tat-derived peptides inhibit cleavage at sites which intact tat does not protect. These results are consistent with transcription studies which have suggested that truncation of tat increases nonspecific binding.

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Selected References

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