Characterization of the 5 S RNA binding activity of Xenopus zinc finger protein p43.

One major component of the Xenopus 42 S ribonucleoprotein (RNP) storage particle is the p43 protein. The 5 S RNA binding protein is structurally similar to TFIIIA, containing nine zinc finger domains. The RNA binding properties of recombinant p43 were characterized using a nitrocellulose filter binding assay. The experimental conditions necessary for in vitro p43-5 S RNA complex formation include: pH 7.5, 0.1 M KCl and incubation at 22 degrees C. Under these conditions, the protein binds to Xenopus oocyte 5 S RNA with an apparent association constant of 1.61(+/- 0.12) x 10(9) M-1. A series of mutations in 5 S RNA were used to determine which sequence and structural features of the 5 S RNA are required for high affinity binding of p43. The primary contact points for p43 include the sequences and structures of stems II, V and loop D of the 5 S RNA. Although p43 and TFIIIA are structurally similar and are both relatively insensitive to mutations in the 5 S RNA, they do require different features of the 5 S RNA molecule for high affinity binding.