RNA RECOGNITION BY THE JOINT ACTION OF 2 NUCLEOLIN RNA-BINDING DOMAINS - GENETIC-ANALYSIS AND STRUCTURAL MODELING

The interaction of nucleolin with a short stem-loop structure (NRE) re quires two contiguous RNA-binding domains (RED 1+2). The structural ba sis for RNA recognition by these RBDs was studied using a genetic syst em in Escherichia coli. Within each of the two domains, we identified several mutations that severely impair interaction with the RNA target . Mutations that alter RNA-binding specificity were also isolated, sug gesting the identity of specific contacts between RED 1+2 amino acids and nucleotides within the NRE stem-loop. Our data indicate that both RBDs participate in a joint interaction with the NRE and that each dom ain uses a different surface to contact the RNA. The constraints provi ded by these genetic data and previous mutational studies have enabled us to propose a three-dimensional model of nucleolin RED 1+2 bound to the NRE stem-loop.