The carboxyterminal zinc fingers of TFIIIA interact with the tip of helix V of 5S RNA in the 7S ribonucleoprotein particle.
Immature Xenopus laevis oocytes contain large quantities of a 7S ribonucleoprotein particle containing transcription factor IIIA (TFIIIA) and 5S RNA in a 1:1 molar ratio. We have reconstituted RNPs containing 5S RNA and either intact TFIIIA or proteolytic fragments that represent progressive C-terminal deletions of the protein. A partial trypsin digestion fragment encompassing the amino terminal seven zinc fingers of TFIIIA rebinds 5S RNA with nearly the same affinity as intact TFIIIA. We have compared the RNase protection patterns resulting from binding of intact and deleted forms of TFIIIA. RNAse protection assays using cobra venom nuclease were performed on complexes reconstituted with 5' and 3' end-labeled 5S RNA. Similar experiments with 3' end-labeled 5S RNA were performed with nuclease alpha-sarcin. With both nucleases, nucleotides in helix V of 5S RNA show more complete protection from nuclease cleavage when the RNA is bound to intact TFIIIA than when it is bound to a 20 kDa tryptic fragment of TFIIIA lacking the C-terminal portion of the protein. These results suggest that fingers 8 and 9 of TFIIIA interact with the distal portion of helix V in the 5S RNA.
Sands
M S
MS
Department of Pharmacology, State University of New York, Stony Brook 11794.
Bogenhagen
D F
DF
D000818:Animals; D001483:Base Sequence; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010446:Peptide Fragments / Q000378:metabolism; D012341:RNA, Ribosomal, 5S / Q000378:metabolism*; D012261:Ribonucleoproteins / Q000235:genetics*; D017411:Ribonucleoproteins, Small Nuclear; D035561:Transcription Factor TFIIA; D014157:Transcription Factors / Q000378:metabolism*; D014357:Trypsin / Q000378:metabolism; D014982:Xenopus laevis; D016335:Zinc Fingers
D010446:Peptide Fragments; D012341:RNA, Ribosomal, 5S; D012261:Ribonucleoproteins; D017411:Ribonucleoproteins, Small Nuclear; D035561:Transcription Factor TFIIA; D014157:Transcription Factors; D014357:Trypsin