Article
- The EMBO Journal (1997) 16, 5235 - 5246
- doi:10.1093/emboj/16.17.5235
RNA recognition by the joint action of two nucleolin RNA-binding domains: genetic analysis and structural modeling
Philippe Bouvet1, Chaitanya Jain2, Joel G. Belasco2, François Amalric1 and Monique Erard1
- Laboratoire de Biologie Moléculaire Eucaryote, Institut de Biologie Cellulaire et de Génétique du CNRS, UPR 9006, 118 route de Narbonne, 31062 Toulouse Cedex, France
- Skirball Institute of Biomolecular Medicine, New York University Medical Center, 540 First Avenue, New York, NY 10016, USA
Correspondence to:
Philippe Bouvet, E-mail: Bouvet@ibcg.biotoul.fr
Received 9 April 1997; Revised 2 June 1997
Abstract
The interaction of nucleolin with a short stem–loop structure (NRE) requires two contiguous RNA-binding domains (RBD 1+2). The structural basis for RNA recognition by these RBDs was studied using a genetic system 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, suggesting the identity of specific contacts between RBD 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 domain uses a different surface to contact the RNA. The constraints provided by these genetic data and previous mutational studies have enabled us to propose a three-dimensional model of nucleolin RBD 1+2 bound to the NRE stem–loop.
Keywords:
- Escherichia coli genetics,
- nucleolin,
- RNA-binding domain,
- RNA-binding specificity,
- structural modeling
