FUNCTIONAL INTERACTIONS BETWEEN THE ZINC FINGERS OF XENOPUS TRANSCRIPTION FACTOR IIIA DURING 5S RIBOSOMAL-RNA BINDING

We have used a collection of mutant forms of Xenopus transcription fac tor IIIA (TFIIIA) to study its interaction with 5S rRNA. This collecti on includes a set of nine mutant proteins, each of which contains a st ructural disruption in one of the nine zinc fingers of TFIIIA (broken- finger mutants), and a pair of complementary N- and C-terminal truncat ion mutants. Equilibrium and kinetic binding analyses in conjunction w ith RNAse protection and interference assays have been used to charact erize the RNA-protein interaction in each case, We find that alternati ve binding modes are available for specific, high affinity recognition of 5S rRNA by TFIIIA, These binding modes are distinct kinetically an d structurally, and the mode of recognition adopted by wild-type TFIII A when binding to intact 5S rRNA is dependent on the structural integr ity of zinc fingers 5 and 6 in TFIIIA and continuity of the sugar-phos phate backbone in loop A of 5S rRNA. Disruption of any of these compon ents allows adoption of one or more alternative modes of binding. In t he wild-type TFIIIA-5S rRNA complex, some portions of TFIIIA, most not ably the N-terminal three zinc fingers, are prevented from interacting with 5S rRNA in an energetically optimal way, and instead adopt a mod e of binding that represents a compromise with the rest of the protein .