A difference in the importance of bulged nucleotides and their parent base pairs in the binding of transcription factor IIIA to Xenopus 5s RNA and 5s RNA genes. Nucleic Acids Res

Individual bulge loops present in Xenopus 5S RNA (positions 49A-A50 in helix HI, Q53 in helix II and A83 in helix IV), were deleted by site directed mutagenesis. The interaction of these mutant 5S RNA molecules with ihlilA was measured by a direct binding assay and a competition assay. The results of these experiments show that none of the bulged nucleotides in Xenopus 5S RNA are required for the binding of TFITJA. The affinity of the mutant 5S RNA genes for TFIIIA was also studied by a filter binding assay. In contrast to the effect that deleting bulged nucleotides had on the TFHIA-RNA binding affinity, deletion of the corresponding A-T base pair at position +83 in 5S DNA was found to reduce the apparent association constant of TFIIIA by a factor of four-fold