Recognition of pre-formed and flexible elements of an RNA stem-loop by nucleolin

Nucleolin is an abundant nucleolar protein which is essential for ribosome biogenesis. The first two of its four tandem RNA-binding domains (RBD12) sp ecifically recognize a stem-loop structure containing a conserved UCCCGA se quence in the loop called the nucleolin-recognition element (NRE). We have determined the structure of the consensus SELEX NRE (sNRE) by NMR spectrosc opy. In both the free and bound RNA the top part of the stem forms a loop E (or S-turn) motif. In the absence of protein, the structure of the hairpin loop is not well defined due to conformational heterogeneity, and appears to be in equilibrium between two families of conformations. Titrations of R BD1, RBD2, and RBD12 with the sNRE show that specific binding requires RBD1 2. Ln complex with RBD12 the hairpin loop interacts specifically with the p rotein and adopts a well-defined structure which shares some of the feature s of the free form. The loop E motif also has specific interactions with th e protein. Implications of these findings for the mechanism of recognition of RNA structures by modular proteins are discussed. (C) 2001 Academic Pres s.